Pesticidal Nucleic Acids and Proteins and Uses Thereof

Abstract

The invention provides compositions comprising polynucleotide molecules encoding certain pesticidal polypeptides which exhibit plant parasitic nematode and/or insect control properties, and are particularly directed to controlling plant parasitic pest species of nematodes and insects known to infest crop plant species. Methods for controlling pests are disclosed in which the toxic proteins are provided in the diet of the targeted plant pests. The invention also provides compositions such as nucleic acids, proteins, and plant and bacterial cells, plants, and seeds containing the nucleic acid and protein compositions, as well as methods and kits for identifying, detecting, and isolating the compositions of the present invention. The invention further provides a method of producing crops from recombinant seeds which contain the polynucleotide molecules encoding the pesticidal polypeptides of the present invention.

Description

CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Application Ser. No. 61/441,697 and U.S. Provisional Application Ser. No. 61/441,709, both filed Feb. 11, 2011, which are incorporated herein by reference.

INCORPORATION OF SEQUENCE LISTING

[0002] The Sequence Listing accompanying this application is contained within the computer readable file "38-21(57560) SEQUENCE LISTING_ST25.txt" submitted electronically and contemporaneously with the filing of this application through the USPTO EFS-Web. The file is 105 kilobytes (measured in MS-Windows), was created on 19 JAN 2012, and is incorporated herein by reference.

FIELD OF THE INVENTION

[0003] The invention relates to novel polynucleotide and protein compositions that, when expressed and or produced in plants, impart resistance to plant pathogenic nematodes and insect infestation. The polynucleotides and proteins can be expressed in plant and bacterial cells, and the plant cells can be regenerated into transgenic (recombinant) plants, plant tissues, plant parts, and seeds. Compositions derived from such plants, plant materials, and seed that contain detectable amounts of such polynucleotides and proteins are included within the scope of the invention. The invention also relates to compositions and methods for controlling plant pathogenic nematodes and insect pests of crop plants.

BACKGROUND OF THE INVENTION

[0004] The increasing human population will require higher yields of food, feed, and fiber from crop plants on decreasing amounts of arable land. Several types of insects and nematodes are known to reduce yield of crops produced from plants. Plant pests damage plant parts, including roots, developing flower buds, flowers, leaves, stems, and seeds, which leads to lower yields.

[0005] Traditional approaches for controlling plant pests have used chemical control agents and construction of inter-specific hybrids between crops and their wild-type relatives as sources of resistant germplasm. Chemical pest control agents, although effective, have several disadvantages. Many chemical control agents are expensive to manufacture, and are characterized as pollutants because they persist in the environment as a result of their resistance to microbial degradation. Chemical control agents require on-farm formulation, which increases the safety risk to the farmer due to the exposure to chemical agent formulations. The chemical agent formulations have to be applied at least once and often, more than once per growing season, increasing the carbon footprint related to these compositions. Methods and compositions employing plant biotechnology pest control agents are also effective means for controlling plant pests, for instance through plant expression of one or more pest control agents that are generally selectively toxic to a particular target pest when ingested by the pest. Unlike chemical agents, biotech approaches have been demonstrated to be environmentally friendly, have no known safety risks when used by farmers, and are economical in terms of carbon footprint impact and ease of use for deployment by the farmer. However, there are only a few examples of such biotech compositions and methods for controlling such pests, and even fewer if any examples of any biotechnology approaches that have demonstrated efficacy in controlling plant pathogenic nematodes. Thus, there is a need for new compositions and methods for protecting plants from such pest infestation, generally for the purpose of maintaining and enhancing yields of crops produced from such plants, and for sustaining and providing food, feed and fiber for the increasing human population.

SUMMARY OF THE INVENTION

[0006] Polynucleotide molecules are provided encoding exemplary pesticidal polypeptides as set forth in SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60. Polypeptides having an amino acid sequence exhibiting from at least about 45% to about 99.9% identity to the pesticidal protein (polypeptide) sequences as set forth in any of the foregoing protein sequences (any percentage in between 45 and 99.9) and exhibiting substantially equivalent (biologically functional equivalent) pesticidal activity as any one of these sequences are specifically contemplated. Fragments of these polypeptide sequences that exhibit the requisite pesticidal activity are intended to be within the scope of the present invention. Such polynucleotides may be extracted and/or obtained directly from a host cell or made artificially through various means of synthesis, and in either case, are considered to be recombinant polynucleotides.

[0007] Polynucleotides containing one or more nucleotide sequence segments encoding the pesticidal proteins of the present invention are provided, which may be operably linked to a heterologous promoter that initiates expression of the sequence region in a designated host cell, resulting in the production or manufacture of the pesticidal protein in the host cell. The promoter may include a plant-expressible promoter, a promoter that functions in one or more species of bacteria, and a yeast functional promoter, or combinations thereof. The plant-expressible promoter may include any number of promoters known in the art, including but not limited to corn sucrose synthetase 1 promoter, corn alcohol dehydrogenase 1 promoter, corn light harvesting complex promoter, corn heat shock protein promoter, pea small subunit RuBP carboxylase promoter, Ti plasmid mannopine synthase promoter, Ti plasmid nopaline synthase promoter, petunia chalcone isomerase promoter, bean glycine rich protein 1 promoter, Potato patatin promoter, lectin promoter, CaMV 35S promoter, FMV promoter, ubiquitin promoters promoter, and the S E9 small subunit RuBP carboxylase promoter.

[0008] Isolated polynucleotide segments are provided for use as probes and/or primers, which may be from about 20 to about 1000 contiguous nucleotides in length or any length in between twenty and one thousand contiguous nucleotides, and exhibit at least about 90% identity to the same contiguous length of nucleotides as set forth in any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:25, SEQ ID NO:27, SEQ ID NO:29, SEQ ID NO:31, SEQ ID NO:33, SEQ ID NO:35, SEQ ID NO:37, SEQ ID NO:39, SEQ ID NO:41, SEQ ID NO:43, SEQ ID NO:45, SEQ ID NO:47, SEQ ID NO:49, SEQ ID NO:51, SEQ ID NO:53, SEQ ID NO:55, SEQ ID NO:57, SEQ ID NO:59, SEQ ID NO:61, and SEQ ID NO:63, or the complement of any of the foregoing polynucleotide sequences.

[0009] In another aspect of the invention, polynucleotides encoding any of the pesticidal polypeptides set forth above are provided in recombinant expression cassettes. The expression cassettes can be provided in vectors for use in replicating, maintaining and transferring the nucleic acid component encoding the pesticidal proteins of the present invention. The vectors of the present invention contain at least a sequence region that encodes the polypeptide as set forth above. The vector includes a plasmid, baculovirus, artificial chromosome, virion, cosmid, phagemid, phage, or viral vector.

[0010] Host cells may be any appropriate transgenic host cell including but not limited to microbial cells (microorganisms) such as an Agrobacterium, a Bacillus, an Escherichia, a Salmonella, a Pseudomonas, a Rhizobium bacterial cell, a yeast cell such as a pichia yeast or saccharomyces species yeast cell, or a plant cell. Vectors as described above can be provided in a transgenic microbial host cell. The transgenic microbial host cell includes a prokaryotic or eukaryotic host cell. The transgenic prokaryotic host cell is a bacterial cell and the transgenic eukaryotic host cell is a plant or a fungal/yeast cell. The transgenic bacterial cell includes a recombinant bacterium including a Bacillus thuringiensis, Bacillus subtilis, Bacillus megaterium, Bacillus cereus, Bacillus laterosperous, Escherichia, Salmonella, Agrobacterium, Rhizobium, or Pseudomonas cell. The transgenic plant host cell includes a monocotyledonous or dicotyledonous plant cell and may include any plant cell from the Group of Plants or Plant Group set forth below. To the extent that a microbial cell is a plant cell, the cell can be obtained from any plant, plant tissue, plant part or seed from a plant selected from the group consisting of any of the following, including but not limited to barley, bean, broccoli, cabbage, canola (rapeseed), carrot, cassava, castor, cauliflower, celery, chickpea, Chinese cabbage, coffee, corn (including sweet corn), clover, cotton, a cucurbit, cucumber, deciduous trees (including but not limited to banana, citrus, eucalyptus, nut trees (including but not limited to hickory, pecan, and walnut trees), oak trees (including but not limited to live oak, pin oak, and post oak trees), olive, palm (including coconut palm), poplar, sweet gum, and rootstocks of all of the preceding trees), eggplant, evergreen trees (including but not limited to Douglas fir), flax, garlic, grape, grasses (including but not limited to alfalfa, pasture grass, switchgrass, and turf grass), hops, leek, lettuce, millets, melons (including but not limited to cantaloupe, honeydew melon, and watermelon), oat, onion, pea, peanut, pepper, pigeonpea, pine (including Loblolly pine, Radiata pine, and Southern pine), potato, pumpkin, radish, rice, rye, safflower, shrub, sorghum, soybean, spinach, squash, strawberry, sugar beet, sugarcane, sunflower, sweet corn, sweet potato, tea, tobacco, tomato, triticale, or wheat. The aforementioned are referenced herein as the "Group of Plants" or the "Plant Group".

[0011] Recombinant plants, plant tissue, plant parts, or seed contain the polynucleotides of the present invention and express the proteins of the present invention from such polynucleotides. The plant part is a leaf, a stem a flower, a sepal, a fruit, a root, or a seed. Products produced from a recombinant plant of the present invention are also contemplated, and can include at least any of the following: oil, meal, lint and seed of the recombinant plant. The polynucleotides and proteins of the present invention are present in a detectable amount in the plants and plant products, and are useful at least as markers for tracking the presence of seeds and plant tissues containing the polynucleotide and proteins through trade and commerce, in fields of crops, and in various embodiments referenced herein.

[0012] There is provided a method of detecting and/or isolating in or from a biological sample, a polynucleotide molecule encoding a pesticidal polypeptide of the present invention in which the steps of the method include (i) selecting a pair of oligonucleotide primers that produce an amplicon encoding all or a representative amount of the pesticidal polypeptide of the present invention when used together in an amplification reaction with the biological sample containing the polynucleotide; (ii) producing the amplicon from the polynucleotide; (iii) detecting and/or isolating the amplicon; and (iv) generating nucleotide sequence information corresponding to the amplicon to identify and confirm the presence (or absence) of a segment of a polynucleotide molecule encoding all or a representative amount of the pesticidal polypeptide. Alternatively, the detecting and/or isolating step can be conducted by providing a polynucleotide probe derived from a sufficient length of DNA or RNA encoding the pesticidal polypeptide that hybridizes under specific or under stringent hybridization conditions to such a polynucleotide encoding a pesticidal polypeptide of the present invention.

[0013] Methods of controlling or killing a target lepidopteran pest, coleopteran pest, or plant pathogenic nematode pest population are provided and include contacting the pest population with a pesticidally-effective amount of the polypeptide as set forth above. The "lepidopteran pest population" includes Spodoptera frugiperda, Spodoptera exigua), Mamestra configurata, Agrotis ipsilon, Trichoplusia ni, Pseudoplusia includens, Anticarsia gemmatalis, Hypena scabra, Heliothis virescens, Agrotis subterranea, Pseudaletia unipuncta, Agrotis orthogonia, Ostrinia nubilalis, Amyelois transitella Crambus caliginosellus, Herpetogramma licarsisalis, Homoeosoma electellum, Elasmopalpus lignosellu, Cydia pomonella, Endopiza viteana, Grapholita molesta, Suleima helianthana, Plutella xylostella, Pectinophora gossypiella, Lymantria dispar, Blatta orientalis, Blatella asahinai, Blattella germanica, Supella longipalpa, Periplaneta americana, Periplaneta brunnea, Leucophaea maderae, Alabama argillacea, Archips argyrospila, A. rosana, Chilo suppressalis, Cnaphalocrocis medinalis, Crambus caliginosellus, C. teterrellus, Diatraea grandiosella, D. saccharalis, Earias insulana, E. vittella, Helicoverpa armigera, H. zea, Heliothis virescens, Herpetogramma licarsisalis, Lobesia botrana, Pectinophora gossypiella, Phyllocnistis citrella, Pieris brassicae, P. rapae, Plutella xylostella, Spodoptera exigua, S. litura, S. frugiperda, Tuta absoluta. The "coleopteran pest population" includes Anthonomus grandis, Lissorhoptrus oryzophilu, Sitophilus granaries, Sitophilus oryzae, Hypera punctata, Sphenophorus maidis, Leptinotarsa decemlineata, Diabrotica virgifera virgifera, Diabrotica barberi, Diabrotica undecimpunctata howardi, Chaetocnema pulicaria, Phyllotreta cruciferae, Colaspis brunnea, Oulema melanopus, Zygogramma exclamationis, Epilachna varivestis, Popillia japonica, Cyclocephala boreali, Cyclocephala immaculata, Rhizotrogus majalis, Phyllophaga crinita, Ligyrus gibbosus, Melanotus spp., Conoderus spp., Limonius spp., Agriotes spp., Ctenicera spp., Aeolus spp., Eleodes spp. The "plant pathogenic nematode population" includes Heterodera glycines (soybean cyst nematode), Heterodera schachtii (beet cyst nematode), Heterodera avenae, Globodera rostochiensis, Globodera pailida, Pratylenchus zeae (a root knot nematode), Meloidogyne javanica, Pratylenchus brachyurus (a root knot nematode), Meloidogyne hapla, Meloidogyne incognita.

[0014] An alternative method for controlling such plant pest infection includes providing a pest inhibitory amount of a pesticidal polypeptide of the present invention to a pest susceptible to the polypeptide, thereby controlling the pest. The pest is an insect or a nematode. The insect may be any insect within the taxonomical orders including Coleoptera, Diptera, Hymenoptera, Lepidoptera, Mallophaga, Homoptera, Hemiptera, Orthoptera, Thysanoptera, Dermaptera, Isoptera, Anoplura, Siphonaptera, or Trichoptera (hereinafter, the "Insect Orders"). The nematode may be from any genus of nematodes referred to as Acontylus, Anguina, Aorolaimus, Aphasmatylenchus, Aphelenchoides, Aphelenchus, Atalodera, Atylenchus, Bakernema, Belonolaimus, Brachydrorus, Bursaphelenchus, Cacopaurus, Caloosia, Carphodorus, Criconema, Criconemella, Cryphodera, Ditylenchus, Dolichodorus, Eutylenchus, Globodera, Gracilacus, Helicotylenchus, Hemicriconemoides, Hemicycliophora, Heterodera, Hirschmanniella, Histotylenchus, Hoplolaimus, Hoplotylus, Longidorus, Macrotrophurus, Meloidodera, Meloidogyne, Merlinius, Morulaimus, Nacobbus, Nothanguina, Nothotylenchus, Paralongidorus, Paratrichodorus, Paratrophurus, Paratylenchus, Peltamigratus, Pratylenchoides, Pratylenchus, Psilenchus, Radopholoides, Radopholus, Rhadinaphelenchus, Rototylenchus, Rotylenchoides, Rotylenchus, Sarisodera, Scutellonema, Sphaeronema, Subanguina, Telotylenchoides, Telotylenchus, Trichotylenchus, Trophonema, Trophotylenculus, Trophurus, Tylenchorhynchus, Tylenchulus, Tylenchus, Tylodorus, Xiphinema, or Zygotylenchus (hereinafter, the "Nematode Species"). In related embodiments, the nematode species includes cyst and related nematodes such as Heterodera glycines (soybean cyst nematode), Heterodera schachtii (beet cyst nematode), Heterodera avenae (cereal cyst nematode), and Globodera rostochiensis and Globodera pailida (potato cyst nematodes), Pratylenchus zeae, Meloidogyne javanica, Pratylenchus brachyurus, Meloidogyne hapla, or Meloidogyne incognita (hereinafter, the "Cyst Nematode" group). The pest inhibitory amount of the pesticidal polypeptide is provided in the diet of the pest, and the diet of the pest can be a part of a recombinant plant, seed of such plant, or product of the plant. The pest inhibitory amount of the polypeptide may also be provided in a topical formulation to a plant. Such formulation could include a preparation containing bacterial cells, bacterial spores, and parasporal crystals which contain or are producing one or more of the polypeptides/toxic agents of the present invention in a sufficient amount to inhibit the pest infestation of the plant to which the formulation is applied. A formulation for controlling nematode or insect species within the scope of the present invention may consist of recombinant bacterial cells and/or spores which may be producing the toxic proteins of the present invention, or parasporal crystals that contain pesticidal amounts of the polypeptide. The bacterial cells, spores, or parasporal crystals are typically from Bacillus species. Antibodies are contemplated that specifically bind to a polypeptide having the amino acid sequence as set forth in any of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and/or SEQ ID NO:60, or a peptide, or an epitope derived therefrom. Particularly, purified antibodies that specifically bind to one or more of the polypeptides of the present invention, or to a peptide or epitope derived from the proteins of the present invention are contemplated.

[0015] Such antibodies are useful at least in methods of detecting pesticidal polypeptides such as those set forth in SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and/or SEQ ID NO:60 in a biological sample. A method of detecting such proteins could include the steps of contacting the biological sample with an antibody that specifically binds to one or more of the proteins of the present invention, and detecting the binding of the antibody to the pesticidal polypeptide. Alternatively, proteins of the present invention, or proteins that are substantially related to the proteins of the present invention can be detected in or isolated from a biological sample either by directly identifying the protein in the sample using for example, antibodies as indicated above, or by screening for the presence of a polynucleotide encoding the pesticidal protein. Detecting the polynucleotide encoding such protein could include the steps of: i) selecting a pair of primers that produce an amplicon encoding the pesticidal protein when used together in an amplification reaction with the polynucleotide; ii) producing the amplicon by using the polynucleotide as a template in the amplification reaction; iii) detecting/isolating the amplicon; iv) generating DNA sequence information corresponding to the amplicon to confirm that the amplicon encodes the pesticidal protein; and v) testing the pesticidal protein to confirm pesticidal activity. Alternatively, a method for detecting the protein of the present invention, or a related pesticidal protein such as a δ-endotoxin polypeptide, in a biological sample could include the steps of: i) obtaining a biological sample suspected of containing a δ-endotoxin polypeptide; ii) contacting the sample with an antibody that specifically binds to the polypeptide under conditions effective to allow the formation of immune complexes; and iii) detecting the immune complexes so formed. Another alternative method for detecting a target pesticidal polypeptide of the present invention in a sample may include the steps of: i) contacting the sample with an antibody that specifically binds the target pesticidal polypeptide; ii) detecting the binding of the antibody to the target in the sample; and iii) identifying the target as a pesticidal polypeptide exhibiting at least 90% amino acid sequence identity to any one of the proteins set forth in SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and/or SEQ ID NO:60.

[0016] Detection methods can be conducted using reagents and instructions packaged together in kit form and are useful for detecting the proteins and polynucleotides of the present invention. Such kits could include a first reagent or antibody that binds specifically to the polypeptide, or specifically to a peptide or an epitope derived therefrom; and a second reagent such as a control polypeptide corresponding to any of the proteins as set forth in any of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and/or SEQ ID NO:60, or a peptide, or an epitope derived therefrom.

[0017] In another aspect of the present invention, there is provided a method of preparing insect resistant plants. Such plants can be prepared by contacting a recipient plant cell with a transgene that encodes one or more of the polypeptides of the present invention under conditions permitting the uptake of the transgene by the cell, and selecting a recipient cell in which the transgene has been incorporated into the cell genome, and regenerating a plant from the selected recipient cell. The regenerated plant is confirmed to be a fertile transgenic plant exhibiting pest resistance, and the pest resistance includes resistance to plant pathogenic nematode infestation and one other pest resistance selected from resistance against to a coleopteran insect or to a lepidopteran insect. The contacting step includes any one or ore of the methods known in the art, including microprojectile bombardment, electroporation or Agrobacterium-mediated plant cell transformation. The regenerated plant is resistant to at least one of the members of the plant parasitic nematode group including Heterodera species, Globodera species, Meloidogyne species, Rotylenchulus species, Hoplolaimus species, Belonolaimus species, Pratylenchus species, Longidorus species, Paratrichodorus species, Ditylenchus species, Xiphinema species, Dolichodorus species, Helicotylenchus species, Radopholus species, Hirschmanniella species, Tylenchorhynchus species, or Trichodorus species.

[0018] Transgenic seed containing one or more polynucleotide segments encoding one or more of the proteins of the present invention may be produced comprising the steps of: transforming a plant with a transgene that encodes the polypeptide as set forth above, the transgene operably linked to a promoter that expresses the transgene in a plant, thereby obtaining a fertile transgenic plant comprising the transgene; and growing the plant under appropriate conditions to produce the transgenic seed.

[0019] Progeny of any generation of a pest resistance-enhanced fertile transgenic plant can be produced from such transgenic plants and seeds of the foregoing plants and seed, wherein the progeny contain the polynucleotide and encode the protein(s) of the present invention, and has enhanced pest resistance against a coleopteran insect, lepidopteran insect, or a plant pathogenic nematode relative to the corresponding non-transgenic plant.

[0020] Pest resistant plants can be produced by following the method of: (a) crossing a pest resistant plant comprising a transgene that encodes the polypeptide as set forth above with another plant; (b) obtaining at least one progeny plant derived from the cross of (a); and (c) selecting progeny that comprises the transgene, wherein the progeny is resistant against a coleopteran insect, lepidopteran insect, or a plant pathogenic nematode.

[0021] Seed can be produced from the plants of the present invention. Seed containing a polynucleotide molecule encoding one or more of the proteins of the present invention, whether homogyzous or heterozygous for the particular transgenic allele, can be packaged for planting in a field, and a crop can be produced from the planted seed. The crop from such plants can be harvested, and if seed of the harvested generation are the crop (such as soybean, rice, wheat, canola or corn and the like), at least 50% of the harvested crop are seed containing the polynucleotide molecule.

[0022] Commodity products (or biological samples) containing a plant or plant part as set forth above that can be shown to contain a detectable amount of a polypeptide having the amino acid sequence of any of the proteins of the present invention, or polynucleotides encoding any such protein. The detection of the polypeptide or the polynucleotide in the commodity (or biological sample) is determinative of the presence of the plant or plant part in the commodity (or biological sample), and all such commodity products in which the polypeptide is detectable to a level of at least about (i) one part per million, (ii) or one nanogram per gram fresh weight of tissue, are within the scope of the present invention. A plant cell of the present invention may be regenerated into a recombinant plant which can produce a plant part containing any of the proteins of the present invention. The plant part includes a leaf, a stem a flower, a sepal, a fruit, a root, or a seed. Products produced from a recombinant plant or plant part contain a detectable amount of any one of the proteins of the present invention, or polynucleotide segments encoding such proteins. Such products include oil, meal, lint and seed of such recombinant plants. The detectable amount of the proteins and/or polynucleotides are useful as molecular markers for tracking and/or identifying the presence of seeds and plant tissues of the present invention as these are moved through commerce.

[0023] The proteins of the present invention originate from Bacillus thuringiensis species of bacteria, and as such, are likely to be characterized as delta-endotoxins, and are typically produced from a recombinant polynucleotide. Such delta endotoxin proteins will have an amino acid sequence that exhibits at least from about 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, 99.9%, or 100% amino acid sequence identity to the amino acid sequence as set forth in any of the sequences shown in SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60. Each such protein will preferably include at least 50, or from about 50 to about 100, or from about 50 to about 300 contiguous amino acids present in any full length protein sequence set forth in the sequences referenced above, and the toxin proteins are preferably encoded by a polynucleotide segment that hybridizes under stringent conditions to the polynucleotide coding sequences as set forth in any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:25, SEQ ID NO:27, SEQ ID NO:29, SEQ ID NO:31, SEQ ID NO:33, SEQ ID NO:35, SEQ ID NO:37, SEQ ID NO:39, SEQ ID NO:41, SEQ ID NO:43, SEQ ID NO:45, SEQ ID NO:47, SEQ ID NO:49, SEQ ID NO:51, SEQ ID NO:53, SEQ ID NO:55, SEQ ID NO:57, SEQ ID NO:59, SEQ ID NO:61, and SEQ ID NO:63.

[0024] Compositions containing the proteins of the present invention are provided in an agriculturally-acceptable carrier. The composition may contain a recombinant Bacillus thuringiensis cell extract, cell suspension, cell homogenate, cell lysate, cell supernatant, cell filtrate, or cell pellet in which at least a pest inhibitory amount of one or more of the proteins of the present invention are provided, and the composition can be provided in the form of a powder, dust, pellet, granule, spray, emulsion, colloid, or solution. The composition may be prepared by desiccation, lyophilization, homogenization, extraction, filtration, centrifugation, sedimentation, or concentration of a culture of recombinant Bacillus thuringiensis cells or spore crystals containing one or more of the proteins of the present invention. The pesticidal composition preferably contains from about 1% to about 99% by weight of one or more of the pesticidal proteins described herein.

[0025] The proteins of the present invention can be obtained in substantially concentrated and/or purified form by a process which may include the steps of i) culturing recombinant Bacillus thuringiensis cells containing one or more recombinant polynucleotide as set forth above under conditions effective to produce the pesticidal protein, and obtaining the pesticidal polypeptide so produced. The polypeptide will preferably contain the contiguous amino acid sequence as set forth in any of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60.

BRIEF DESCRIPTION OF THE SEQUENCES

[0026] SEQ ID NO 1 represents a native Bt nucleotide sequence encoding a ET34 protein. SEQ ID NO 2 represents an amino acid sequence translation of SEQ ID NO 1. SEQ ID NO 3 represents an artificial sequence encoding a ET34 protein. SEQ ID NO 4 represents an amino acid sequence translation of SEQ ID NO 3 from nucleotide position 1 through nucleotide position 378. SEQ ID NO 5 represents a nucleotide sequence encoding a P139 secretion signal peptide (nucleotide position 1-75) fused in frame to a native Bt nucleotide sequence encoding a ET34 protein (nucleotide position 76-450). SEQ ID NO 6 represents the amino acid sequence translation of SEQ ID NO 5. SEQ ID NO 7 represents a nucleotide sequence encoding a P139 secretion signal peptide (nucleotide position 1-75) fused in frame to a synthetic nucleotide sequence encoding a ET34 protein (nucleotide position 76-450). SEQ ID NO 8 represents the amino acid sequence translation of SEQ ID NO 7. SEQ ID NO 9 represents the native Bt nucleotide sequence encoding a TIC1506 protein. SEQ ID NO 10 represents the amino acid sequence translation of SEQ ID NO 9. SEQ ID NO 11 represents an artificial nucleotide sequence encoding a TIC1506 protein. SEQ ID NO 12 represents the amino acid sequence translation of SEQ ID NO 11. SEQ ID NO 13 represents the native Bt nucleotide sequence encoding a TIC1501 protein. SEQ ID NO 14 represents the amino acid sequence translation of SEQ ID NO 13. SEQ ID NO 15 represents an artificial nucleotide sequence encoding a TIC1501 protein. SEQ ID NO 16 represents the amino acid sequence translation of SEQ ID NO 15. SEQ ID NO 17 represents the native Bt nucleotide sequence encoding a TIC1503 protein. SEQ ID NO 18 represents the amino acid sequence translation of SEQ ID NO 17. SEQ ID NO 19 represents an artificial nucleotide sequence encoding a TIC1503 protein. SEQ ID NO 20 represents the amino acid sequence translation of SEQ ID NO 19. SEQ ID NO 21 represents a native Bt nucleotide sequence encoding a TIC614 protein. SEQ ID NO 22 represents an amino acid sequence translation of SEQ ID NO 21. SEQ ID NO 23 represents an artificial nucleotide sequence encoding a TIC614 protein. SEQ ID NO 24 represents an amino acid sequence translation of SEQ ID NO 23. SEQ ID NO 25 represents a nucleotide sequence encoding a TIC615 protein. SEQ ID NO 26 represents the amino acid sequence translation of SEQ ID NO 25. SEQ ID NO 27 represents an artificial nucleotide sequence encoding a TIC615 protein. SEQ ID NO 28 represents the amino acid sequence translation of SEQ ID NO 27. SEQ ID NO 29 represents the native Bt nucleotide sequence encoding a TIC1277 protein. SEQ ID NO 30 represents the amino acid sequence translation of SEQ ID NO 29. SEQ ID NO 31 represents an artificial nucleotide sequence encoding a TIC1277 protein. SEQ ID NO 32 represents the amino acid sequence translation of SEQ ID NO 31. SEQ ID NO 33 represents the native Bt nucleotide sequence encoding a TIC TIC1278 protein. SEQ ID NO 34 represents the amino acid sequence translation of SEQ ID NO 33. SEQ ID NO 35 represents an artificial nucleotide sequence encoding a TIC TIC1278 protein. SEQ ID NO 36 represents the amino acid sequence translation of SEQ ID NO 35. SEQ ID NO 37 represents the native Bt nucleotide sequence encoding a TIC TIC1310 protein. SEQ ID NO 38 represents the amino acid sequence translation of SEQ ID NO 37. SEQ ID NO 39 represents an artificial nucleotide sequence encoding a TIC1310 protein. SEQ ID NO 40 represents the amino acid sequence translation of SEQ ID NO 39. SEQ ID NO 41 represents the native Bt nucleotide sequence encoding a TIC TIC1311protein. SEQ ID NO 42 represents the amino acid sequence translation of SEQ ID NO 41. SEQ ID NO 43 represents an artificial nucleotide sequence encoding a TIC1311 protein. SEQ ID NO 44 represents the amino acid sequence translation of SEQ ID NO 43. SEQ ID NO 45 represents the native Bt nucleotide sequence encoding a TIC1324 protein. SEQ ID NO 46 represents the amino acid sequence translation of SEQ ID NO 45. SEQ ID NO 47 represents an artificial nucleotide sequence encoding a TIC1324 protein. SEQ ID NO 48 represents the amino acid sequence translation of SEQ ID NO 47. SEQ ID NO 49 represents the native Bt nucleotide sequence encoding a TIC1407 protein. SEQ ID NO 50 represents the amino acid sequence translation of SEQ ID NO 49. SEQ ID NO 51 represents an artificial nucleotide sequence encoding a TIC TIC1407 protein. SEQ ID NO 52 represents the amino acid sequence translation of SEQ ID NO 51. SEQ ID NO 53 represents the native Bt nucleotide sequence encoding a TIC TIC1408 protein. SEQ ID NO 54 represents the amino acid sequence translation of SEQ ID NO 53. SEQ ID NO 55 represents an artificial nucleotide sequence encoding a TIC1408 protein. SEQ ID NO 56 represents the amino acid sequence translation of SEQ ID NO 56. SEQ ID NO 57 represents a native Bt nucleotide sequence encoding a TIC1308 protein. SEQ ID NO 58 represents an amino acid sequence translation of SEQ ID NO 57. SEQ ID NO 59 represents a native Bt nucleotide sequence encoding a TIC1442 protein. SEQ ID NO 60 represents an amino acid sequence translation of SEQ ID NO 59. SEQ ID NO 61 represents an artificial nucleotide sequence encoding a TIC1308 protein. SEQ ID NO 62 represents an amino acid sequence translation of SEQ ID NO 61. SEQ ID NO 63 represents an artificial nucleotide sequence encoding a TIC1442 protein. SEQ ID NO 64 represents an amino acid sequence translation of SEQ ID NO 63.

DETAILED DESCRIPTION OF THE INVENTION

[0027] The invention relates to methods and compositions for pest control in plants, in particular nematode and/or insect control. In one aspect, the invention relates to controlling, preventing or treating nematode and/or insect infection in transgenic plants. The method comprises, in one embodiment, generation of transgenic plants containing a recombinant construct and expression of such construct to impart such pest resistance to plants. The recombinant construct may comprise a nucleotide sequence encoding one or more proteins, wherein the sequence is operably linked to a heterologous promoter functional in a plant cell, and to cells transformed with the recombinant construct. Cells comprising (meaning including but not limited to) the recombinant construct may be prokaryotic or eukaryotic. In particular, eukaryotic cells may be plant cells. Plants and seeds derived from such transformed plant cells are also contemplated. The transgenic plants or parts thereof of the present invention, in one embodiment, produce one or more pesticidal proteins derived from Bacillus thuringiensis bacterial strains.

[0028] The present invention provides heterologous molecules that are expressed in the cytoplasm of the host cell, or if used in a eukaryotic cell such as a plant cell, may also be directed into the plastid of the plant to provide production of the toxic protein, and including, but not limited to, nucleotide segments that encode polypeptides such as SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60 having pesticidal activity. In certain embodiments, the polypeptide having pesticidal activity may share at least about 45%, or at least about 50%, or at least about 51-79%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 98%, or at least 99%, or 100% sequence identity, to any one or more amino acid sequence(s) set forth in SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, or SEQ ID NO:60. The function of the encoded polypeptide may also be determined by measuring the efficacy of the presence of the transgene that encodes it in reducing nematode and/or insect infection, growth, reproduction, or symptomology. For instance, a reduction in root galls, cysts, or worm number of 20% or more, 25% or more, 50% or more, 80% or more, or 95% or more, in a transgenic plant comprising a heterologous nucleotide construct encoding any of the proteins of the present invention, relative to a control plant, for instance an otherwise isogenic plant not comprising the heterologous molecule, under similar conditions, indicates the presence of a functional molecule.

[0029] In certain embodiments, a heterologous molecule provided by the present invention that is directed into the plastid of a plant to provide production of a toxin protein of the present invention may share at least from about 60 to about 79%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 98%, or at least 99%, or 100% sequence identity at the nucleotide level with one or more sequence(s) as set forth in SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:5, SEQ ID NO:7, SEQ ID NO:9, SEQ ID NO:11, SEQ ID NO:13, SEQ ID NO:15, SEQ ID NO:17, SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:23, SEQ ID NO:25, SEQ ID NO:27, SEQ ID NO:29, SEQ ID NO:31, SEQ ID NO:33, SEQ ID NO:35, SEQ ID NO:37, SEQ ID NO:39, SEQ ID NO:41, SEQ ID NO:43, SEQ ID NO:45, SEQ ID NO:47, SEQ ID NO:49, SEQ ID NO:51, SEQ ID NO:53, SEQ ID NO:55, SEQ ID NO:57, SEQ ID NO:59, SEQ ID NO:61, and SEQ ID NO:63. Thus, in particular embodiments, the heterologous molecule may comprise a sequence encoding a heterologous chloroplast transit peptide.

[0030] Yet another aspect of the invention provides methods for production and for use of one or more of the proteins of the present invention to control nematode and/or insect infestation. Thus, methods for production of a toxin, for instance in a plant cell, are provided. The toxin may then be applied to soil prior to, during, or subsequent to planting of a crop, in order to control or reduce nematode infestation or symptomatology of crop plants grown in that soil.

[0031] Unless otherwise noted, terms are to be understood according to conventional usage by those of ordinary skill in the relevant art. Definitions of common terms in molecular biology may also be found in Rieger et al., Glossary of Genetics: Classical and Molecular, 5th edition, Springer-Verlag: New York, 1991; and Lewin, Genes V, Oxford University Press: New York, 1994. The nomenclature for DNA bases as set forth at Title 37 of the United States Code of Federal Regulations, Part 1, section 1.822.

[0032] As used herein, a "transgenic plant" is any plant in which one or more, or all, of the cells of the plant include a transgene. A transgene may be integrated within a nuclear genome or organelle genome, or it may be extra-chromosomally replicating DNA. The term "transgene" means a nucleic acid that is partly or entirely heterologous, foreign, to a transgenic microbe, plant, animal, or cell into which it is introduced. Cells that make up various cell and tissue types of plants include but are not limited to seed, root, leaf, shoot, flower, pollen and ovule.

[0033] "Recombinant DNA" is a polynucleotide having a genetically engineered modification introduced through combination of endogenous and/or exogenous molecules in a transcription unit, manipulation via mutagenesis, restriction enzymes, and the like or simply by inserting multiple copies of a native transcription unit. Recombinant DNA may comprise DNA segments obtained from different sources, or DNA segments obtained from the same source, but which have been manipulated to join DNA segments which do not naturally exist in the joined form. An isolated recombinant polynucleotide may exist, for example as a purified molecule, or integrated into a genome, such as a plant cell, or organelle genome or a microbe plasmid or genome. The polynucleotide comprises linked regulatory molecules that cause transcription of an RNA in a plant cell.

[0034] As used herein, "percent identity" means the extent to which two optimally aligned DNA or protein segments are invariant throughout a window of alignment of components, for example nucleotide sequence or amino acid sequence. An "identity fraction" for aligned segments of a test sequence and a reference sequence is the number of identical components that are shared by sequences of the two aligned segments divided by the total number of sequence components in the reference segment over a window of alignment which is the smaller of the full test sequence or the full reference sequence. "Percent identity" ("% identity") is the identity fraction times 100.

[0035] "Expression" means transcription of DNA to produce RNA. The resulting RNA may be without limitation mRNA encoding a protein, antisense RNA, or a double-stranded RNA for use in RNAi technology. Expression also may refer to translation of RNA, i.e. the production of encoded protein from an mRNA.

[0036] As used herein, "promoter" means regulatory DNA molecules for initializing transcription. A "plant promoter" is a promoter capable of initiating transcription in plant cells whether or not its origin is a plant cell. For example it is well known that certain Agrobacterium promoters are functional in plant cells. Thus, plant promoters include promoter DNA obtained from plants, plant viruses (in particular, double stranded DNA viruses) and bacteria such as Agrobacterium and Bradyrhizobium bacteria. Constitutive promoters generally provide transcription in most or all of the cells of a plant. In particular, promoters such as the FMV promoter (FMV, U.S. Pat. No. 6,051,753), the enhanced 35S promoter (E35S, U.S. Pat. No. 5,359,142), rice actin promoter (U.S. Pat. No. 5,641,876), and various chimeric promoters (U.S. Pat. No. 6,660,911) are useful in the present invention. Examples of promoters under developmental control include promoters that preferentially initiate transcription in certain tissues, such as leaves, roots, or seeds. Such promoters are referred to as "tissue-preferred". Promoters that initiate transcription only in certain tissues are referred to as "tissue specific."

[0037] A number of root-specific or root-enhanced promoters or fragments of such that provide enhanced expression in root tissues relative to other plant tissues have been identified and are known in the art (e.g. U.S. Pat. Nos. 5,110,732, 5,837,848, 5,837,876; 5,633,363; 5,459,252; 5,401,836; 7,196,247; 7,232,940; 7,119,254; and 7,078,589). Examples include root-enhanced or root-specific promoters such as the CaMV-derived as-1 promoter or the wheat PDX1 promoter (U.S. Pat. No. 5,023,179), the acid chitinase gene promoter (Samac et al., Plant Mol. Biol. 25:587-596 (1994); the root specific subdomains of the CaMV35S promoter (Lam et al., Proc. Natl. Acad. Sci. (U.S.A.) 86:7890-7894 (1989); the root-enhanced ORF13 promoter from Agrobacterium rhizogenes (Hansen et al., Mol. Gen. Genet. 254:337-343 (1997); the promoter for the tobacco root-specific gene RB7 (U.S. Pat. No. 5,750,386); and the root cell-specific promoters reported by Conkling et al. (Plant Physiol. 93:1203-1211 (1990). Additional examples include RCc2 and RCc3, promoters that direct root-specific gene transcription in rice (Xu et al., Plant Mol. Biol. 27:237, 1995); soybean root-specific glutamine synthetase promoter (Hire et al., Plant Mol. Biol. 20:207-218, 1992); root-specific control element in the GRP 1.8 gene of French bean (Keller and Baumgartner, Plant Cell 3:1051-1061, 1991.); a root-specific promoter of the mannopine synthase (MAS) gene of Agrobacterium tumefaciens (Sanger et al., Plant Mol. Biol. 14:433-443, 1990); and full-length cDNA clone encoding cytosolic glutamine synthetase (GS), which is expressed in roots and root nodules of soybean (Miao et al., Plant Cell 3:11-22, 1991). See also Bogusz et al., Plant Cell 2:633-641, 1990, where two root-specific promoters isolated from hemoglobin genes from the nitrogen-fixing non-legume Parasponia andersonii and the related non-nitrogen-fixing non-legume Trema tomentosa are described. Leach and Aoyagi (1991) describe their analysis of the promoters of the highly expressed rolC and rolD root-inducing genes of Agrobacterium rhizogenes (see Plant Science (Limerick) 79:69-76). Additional root-preferred promoters include the VfENOD-GRP3 gene promoter (Kuster et al., Plant Mol. Biol. 29(4):759-772, 1995); and rolB promoter (Capana et al., Plant Mol. Biol. 25:681-691, 1994). Examples of nematode-induced promoters include, for instance, the TobRB7 promoter (Opperman et al., Science 263:221-223, 1994), and promoters described in U.S. Pat. Nos. 6,262,344, and 7,193,136.

[0038] The term "resistance," or "tolerance" when used in the context of comparing the effectiveness of a transgene in a transgenic plant, refers to the ability of the transgenic plant to maintain a desirable phenotype when exposed to nematode infestation pressures relative to the phenotype presented by a nematode sensitive non-transgenic plant under similar conditions. The level of resistance can be determined by comparing the physical characteristics of the transgenic plant to non-transgenic plants that either have or have not been exposed to nematode and/or insect infection. Exemplary physical characteristics to observe include plant height, an increase in population of plants that have ability to survive nematode or insect challenge (that is, plants that come in contact with a parasitic nematode or insect may have enhanced root growth, enhanced fruit or grain yield, and decreased reproduction of the nematode or insect infesting the plant or crop, or a decrease in the rate of increase if the pest population). The product of expression of the recombinant DNA may be directly toxic to the nematode (nematicidal) or insect (insecticidal), or may affect the mobility, host finding, feeding site establishment, fecundity or have other nematistatic and/or insectic inhibitory effects.

[0039] "Transformed seed" is the seed which has been generated from the transformed plant. A transformed plant contains transformed cells. A transformed cell is a cell that has been altered by the introduction of an exogenous DNA molecule or in the present invention comprises a heterologous DNA encoding one or more of the proteins of the present invention.

[0040] Pests intended to be within the scope of the present invention include the "lepidopteran pest population" such as Spodoptera frugiperda, Spodoptera exigua, Mamestra configurata, Agrotis ipsilon, Trichoplusia ni, Pseudoplusia includens, Anticarsia gemmatalis, Hypena scabra, Heliothis virescens, Agrotis subterranea, Pseudaletia unipuncta, Agrotis orthogonia, Ostrinia nubilalis, Amyelois transitella Crambus caliginosellus, Herpetogramma licarsisalis, Homoeosoma electellum, Elasmopalpus lignosellu, Cydia pomonella, Endopiza viteana, Grapholita molesta, Suleima helianthana, Plutella xylostella, Pectinophora gossypiella, Lymantria dispar, Blatta orientalis, Blatella asahinai, Blattella germanica, Supella longipalpa, Periplaneta americana, Periplaneta brunnea, Leucophaea maderae, Alabama argillacea, Archips argyrospila, A. rosana, Chilo suppressalis, Cnaphalocrocis medinalis, Crambus caliginosellus, C. teterrellus, Diatraea grandiosella, D. saccharalis, Earias insulana, E. vittella, Helicoverpa armigera, H. zea, Heliothis virescens, Herpetogramma licarsisalis, Lobesia botrana, Pectinophora gossypiella, Phyllocnistis citrella, Pieris brassicae, P. rapae, Plutella xylostella, Spodoptera exigua, S. litura, S. frugiperda, and Tuta absoluta. The "coleopteran pest population" includes Anthonomus grandis, Lissorhoptrus oryzophilu, Sitophilus granaries, Sitophilus oryzae, Hypera punctata, Sphenophorus maidis, Leptinotarsa decemlineata, Diabrotica virgifera virgifera, Diabrotica barberi, Diabrotica undecimpunctata howardi, Chaetocnema pulicaria, Phyllotreta cruciferae, Colaspis brunnea, Oulema melanopus, Zygogramma exclamationis, Epilachna varivestis, Popillia japonica, Cyclocephala boreali, Cyclocephala immaculata, Rhizotrogus majalis, Phyllophaga crinita, Ligyrus gibbosus, Melanotus spp., Conoderus spp., Limonius spp., Agriotes spp., Ctenicera spp., and Aeolus spp., Eleodes spp. The "plant pathogenic nematode population" includes plant parasitic species, for example, Heterodera species, Globodera species, Meloidogyne species, Rotylenchulus species, Hoplolaimus species, Belonolaimus species, Pratylenchus species, Longidorus species, Paratrichodorus species, Ditylenchus species, Xiphinema species, Dolichodorus species, Helicotylenchus species, Radopholus species, Hirschmanniella species, Tylenchorhynchus species, and Trichodorus species, and the like, and specifically includes Heterodera glycines (soybean cyst nematode), Heterodera schachtii (beet cyst nematode), Heterodera avenae, Globodera rostochiensis, Globodera pailida, Pratylenchus zeae (a root knot nematode), Meloidogyne javanica, Pratylenchus brachyurus (a root knot nematode), Meloidogyne hapla, and Meloidogyne incognita.

[0041] The present invention provides recombinant DNA constructs comprising a polynucleotide that, when incorporated in a plant cell, imparts to the plant resistance to nematode and/or insect infection or plant disease caused by such infection (also referred to as infestation). Such constructs also typically comprise a promoter operatively linked to said polynucleotide to provide for expression in the plant cells. Other construct components may include additional regulatory molecules, such as 5' leader regions or 3' untranslated regions (such as polyadenylation sites), intron regions, and transit or signal peptides. Such recombinant DNA constructs can be assembled using methods known to those of ordinary skill in the art.

[0042] Recombinant constructs prepared in accordance with the present invention also generally include a 3' untranslated DNA region (UTR) that typically contains a polyadenylation sequence following the polynucleotide coding region. Examples of useful 3' UTRs include but are not limited to those from the nopaline synthase gene of Agrobacterium tumefaciens (nos), a gene encoding the small subunit of a ribulose-1,5-bisphosphate carboxylase-oxygenase (rbcS), and the T7 transcript of Agrobacterium tumefaciens.

[0043] Constructs and vectors may also include a transit peptide for targeting of a protein product, particularly to a chloroplast, leucoplast or other plastid organelle, mitochondria, peroxisome, or vacuole or an extracellular location. For descriptions of the use of chloroplast transit peptides, see U.S. Pat. No. 5,188,642 and U.S. Pat. No. 5,728,925. Many chloroplast-localized proteins are expressed from nuclear genes as precursors and are targeted to the chloroplast by a chloroplast transit peptide (CTP). Examples of other such isolated chloroplast proteins include, but are not limited to those associated with the small subunit (SSU) of ribulose-1,5,-bisphosphate carboxylase, ferredoxin, ferredoxin oxidoreductase, the light-harvesting complex protein I and protein II, thioredoxin F, enolpyruvyl shikimate phosphate synthase (EPSPS) and transit peptides described in U.S. Pat. No. 7,193,133. It has been demonstrated in vivo and in vitro that non-chloroplast proteins may be targeted to the chloroplast by use of protein fusions with a heterologous CTP and that the CTP is sufficient to target a protein to the chloroplast. Incorporation of a suitable chloroplast transit peptide, such as, the Arabidopsis thaliana EPSPS CTP (CTP2, Klee et al., Mol. Gen. Genet. 210:437-442, 1987), and the Petunia hybrida EPSPS CTP (CTP4, della-Cioppa et al., Proc. Natl. Acad. Sci. USA 83:6873-6877, 1986) has been show to target heterologous EPSPS protein sequences to chloroplasts in transgenic plants. The production of glyphosate tolerant plants by expression of a fusion protein comprising an amino-terminal CTP with a glyphosate resistant EPSPS enzyme is well known by those skilled in the art, (U.S. Pat. No. 5,627,061, U.S. Pat. No. 5,633,435, U.S. Pat. No. 5,312,910, EP 0218571, EP 189707, EP 508909, and EP 924299). Those skilled in the art will recognize that various chimeric constructs can be made that utilize the functionality of a CTP to import various pesticidal proteins of the present invention into the plant cell plastid.

[0044] Stable methods for plant transformation include virtually any method by which DNA can be introduced into a cell, such as by direct delivery of DNA (for example, by PEG-mediated transformation of protoplasts, by electroporation, by agitation with silicon carbide fibers, and by acceleration of DNA coated particles), by Agrobacterium-mediated transformation, by viral or other vectors. One preferred method of plant transformation is microprojectile bombardment, for example, as illustrated in U.S. Pat. Nos. 5,015,580 (soy), 5,550,318 (maize), 5,538,880 (maize), 6,153,812 (wheat), 6,160,208 (maize), 6,288,312 (rice) and 6,399,861 (maize), and 6,403,865 (maize).

[0045] Detailed procedures for Agrobacterium-mediated transformation of plants, especially crop plants, include, for example, procedures disclosed in U.S. Pat. Nos. 5,004,863, 5,159,135, 5,518,908, 5,846,797, and 6,624,344 (cotton); 5,416,011, 5,569,834, 5,824,877, 5,914,451 6,384,301, and 7,002,058 (soy); 5,591,616 5,981,840, and 7,060,876 (maize); 5,463,174 and 5,750,871 (Brassica species, including rapeseed and canola), and in U.S. Patent Application Publications 2004/0244075 (maize), 2004/0087030 (cotton) and 2005/0005321 (soybean). Additional procedures for Agrobacterium-mediated transformation are disclosed in WO9506722 (maize). Similar methods have been reported for many plant species, both dicots and monocots, including, among others, peanut (Cheng et al., Plant Cell Rep., 15:653, 1996); asparagus (Bytebier et al., Proc. Natl. Acad. Sci. U.S.A., 84:5345, 1987); barley (Wan and Lemaux, Plant Physiol., 104:37, 1994); rice (Toriyama et al., Bio/Technology, 6:10, 1988; Zhang et al., Plant Cell Rep., 7:379, 1988; wheat (Vasil et al., Bio/Technology, 10:667, 1992; Becker et al., Plant J., 5:299, 1994), alfalfa (Masoud et al., Transgen. Res., 5:313, 1996); Brassica species (Radke et al., Plant Cell Rep., 11:499-505, 1992); and tomato (Sun et al., Plant Cell Physiol., 47:426-431, 2006). Transgenic plant cells and transgenic plants can also be obtained by transformation with other vectors, such as but not limited to viral vectors (for example, tobacco etch virus (TEV), barley stripe mosaic virus (BSMV), and the viruses referenced in Edwardson and Christie, "The Potyvirus Group Monograph No. 16", 1991, Agric. Exp. Station, Univ. of Florida), plasmids, cosmids, YACs (yeast artificial chromosomes), BACs (bacterial artificial chromosomes) or any other suitable cloning vector, when used with an appropriate transformation protocol such as but not limited to bacterial infection (for example, with Agrobacterium as described above), binary bacterial artificial chromosome constructs, direct delivery of DNA (for example, via PEG-mediated transformation, desiccation/inhibition-mediated DNA uptake, electroporation, agitation with silicon carbide fibers, and microprojectile bombardment). It would be clear to one of ordinary skill in the art that various transformation methodologies can be used and modified for production of stable transgenic plants from any number of plant species of interest. For example the construction of stably inherited recombinant DNA constructs and mini-chromosomes can be used as vectors for the construction of transgenic plants (U.S. Pat. No. 7,235,716).

[0046] Plants of the present invention include, but are not limited to, Acacia, alfalfa, aneth, apple, apricot, artichoke, arugula, asparagus, avocado, banana, barley, beans, beet, blackberry, blueberry, broccoli, brussels sprouts, cabbage, canola, cantaloupe, carrot, cassava, cauliflower, celery, cherry, cilantro, citrus, clementine, coffee, corn, cotton, cucumber, Douglas fir, eggplant, endive, escarole, eucalyptus, fennel, figs, forest trees, gourd, grape, grapefruit, honey dew, jicama, kiwifruit, lettuce, leeks, lemon, lime, loblolly pine, mango, melon, mushroom, nut, oat, okra, onion, orange, an ornamental plant, papaya, parsley, pea, peach, peanut, pear, pepper, persimmon, pine, pineapple, plantain, plum, pomegranate, poplar, potato, pumpkin, quince, radiata pine, radicchio, radish, rapeseed, raspberry, rice, rye, sorghum, Southern pine, soybean, spinach, squash, strawberry, sugarbeet, sugarcane, sunflower, sweet potato, sweetgum, tangerine, tea, tobacco, tomato, turf, a vine, watermelon, wheat, yams, and zucchini. Crop plants are defined as plants which are cultivated to produce one or more commercial products. Examples of such crops or crop plants include but are not limited to soybean, canola, rape, cotton (cottonseeds), peanut, sunflower, pigeon pea, chickpea, and the like, and grains such as corn, wheat, rice, oat, millet, and rye, and the like. Rape, rapeseed and canola are used synonymously in the present disclosure.

[0047] Transformation methods to provide transgenic plant cells and transgenic plants containing stably integrated recombinant DNA are preferably practiced in tissue culture on media and in a controlled environment. Recipient cell targets include but are not limited to meristem cells, callus, immature embryos or parts of embryos, gametic cells such as microspores, pollen, sperm, and egg cells. Any cell from which a fertile plant can be regenerated is contemplated as a useful recipient cell for practice of the invention. Callus can be initiated from various tissue sources, including, but not limited to, immature embryos or parts of embryos, seedling apical meristems, microspores, and the like. Those cells which are capable of proliferating as callus can serve as recipient cells for genetic transformation. Practical transformation methods and materials for making transgenic plants of this invention (for example, various media and recipient target cells, transformation of immature embryos, and subsequent regeneration of fertile transgenic plants) are disclosed, for example, in U.S. Pat. Nos. 6,194,636 and 6,232,526 and U.S. Patent Application Publication 2004/0216189.

[0048] In general transformation practice, DNA is introduced into only a small percentage of target cells in any one transformation experiment. Marker genes are generally used to provide an efficient system for identification of those cells that are transformed by a transgenic DNA construct. Preferred marker genes provide selective markers which confer resistance to a selective agent, such as an antibiotic or herbicide. Any of the antibiotics or herbicides to which a plant cell may be resistant can be a useful agent for selection. Potentially transformed cells are exposed to the selective agent. In the population of surviving cells will be those cells where, generally, the resistance-conferring gene is expressed at sufficient levels to permit cell survival in the presence of the selective agent. Cells can be tested further to confirm integration of the recombinant DNA. Commonly used selective marker genes include those conferring resistance to antibiotics such as kanamycin or paromomycin (val), hygromycin B (aph IV), gentamycin (aac3 and aacC4) and glufosinate (bar or pat), glyphosate (EPSPS), and dicamba (dicamba monooxygenase). Examples of useful selective marker genes and selection agents are illustrated in U.S. Pat. Nos. 5,550,318, 5,633,435, 5,780,708, and 6,118,047. Screenable markers or reporters, such as markers that provide an ability to visually identify transformants can also be employed. Non-limiting examples of useful screenable markers include, for example, a gene expressing a protein that produces a detectable color by acting on a chromogenic substrate (for example, beta-glucuronidase, GUS, uidA, or luciferase, luc) or that itself is detectable, such as green fluorescent protein (GFP, gfp) or an immunogenic molecule. Those of skill in the art will recognize that many other useful markers or reporters are available for use.

[0049] The recombinant DNA constructs of the invention can be stacked with other recombinant DNA for imparting additional agronomic traits (such as in the case of transformed plants, traits including but not limited to herbicide resistance, insect resistance, cold germination tolerance, water deficit tolerance, enhanced yield, enhanced quality, fungal, viral, and bacterial disease resistance) for example, by expressing other transgenes. The recombinant DNA constructs of the present invention can also be transformed into plant varieties that carry natural pest or pathogen resistance genes to enhance the efficacy of the resistance phenotype. Constructs for coordinated decrease and/or increase of gene expression are disclosed in U.S. Patent Application Publication 2004/0126845 A1. Seeds of transgenic, fertile plants can be harvested and used to grow progeny generations, including hybrid generations, of transgenic plants of this invention that include the recombinant DNA construct in their genome. Thus, in addition to direct transformation of a plant with a recombinant DNA construct of this invention, transgenic plants of the invention can be prepared by crossing a first plant having the recombinant DNA with a second plant lacking the construct. For example, the recombinant DNA can be introduced into a plant line that is amenable to transformation to produce a transgenic plant, which can be crossed with a second plant line to introgress the recombinant DNA into the resulting progeny. A transgenic plant of the invention can be crossed with a plant line having other recombinant DNA or naturally occurring genetic regions that confers one or more additional trait(s) (such as, but not limited to, herbicide resistance, pest or disease resistance, environmental stress resistance, modified nutrient content, and yield improvement) to produce progeny plants having recombinant DNA that confers both the desired target sequence expression behavior and the additional trait(s). Typically, in such breeding for combining traits the transgenic plant donating the additional trait is a male line and the transgenic plant carrying the base traits is the female line. The progeny of this cross segregate such that some of the plant will carry the DNA for both parental traits and some will carry DNA for one parental trait; such plants can be identified by markers associated with parental recombinant DNA. Progeny plants carrying DNA for both parental traits can be crossed back into the female parent line multiple times, for example, usually 6 to 8 generations, to produce a progeny plant with substantially the same genotype as one original transgenic parental line but for the recombinant DNA of the other transgenic parental line.

[0050] Other proteins and toxic agents can be used together with one or more proteins of the present invention to control plant pathogenic nematode and/or insect infestation and to reduce the likelihood of development of resistance to any single method of control. Such other proteins and toxic agents include but are not limited to, as applicable to either nematode or insect control, methylketone synthase, dsRNA expressed in the cell and targeting for suppression one or more essential, housekeeping, reproductive or developmental gene, other proteins that are known in the art to be toxic to plant pathogenic nematodes or insects such as Cry and VIP proteins (lifesci.sussex.ac.uk/home/Neil_Crickmore/Bt/index.html" on the world wide web, which is properly referenced as Crickmore et. al. (2010) "Bacillus thuringiensis toxin nomenclature"), as well as chemical nematicides used in seed treatments or soil drenches. Topically applied dsRNA methods are also known in the art that can be applied to a plant expressing one or more of the proteins of the present invention. Such topical applications can be effective in causing a systemic effect in the plant that result in nematode or insect control by applying to the plant a dsRNA molecule that targets for regulation a gene in the plant involved in such resistance. All such combinations are within the scope of the present invention.

[0051] The transgenic plant, plant part, seed or progeny plants of the present invention can be processed into products useful in commerce. These products, commodity products, include but are not limited to meal, flour, oil, hay, starch, juice, protein extract, and fiber.

[0052] The proteins of the present invention have been identified using a variety of methods. One method has been to identify previously known Bt proteins that exhibit a mass less than about 40 kDa, or less than about 35 kDa, or less than about 30 kDa, or less than about 28 kDa, or less than about 25 kDa, or less than about 20 kDa, or less than about 15 kDa. Such proteins include but are not limited to the smaller component of most known binary Bt toxins, such as Cry34/35 (PS149B1), TIC100/101, ET33/34, ET80/76, and the like. Other proteins known in the art include TIC901, TIC1201, TIC407, TIC417, TIC431, ET70, VIP proteins such as VIP3Aa and the like, which are all generally small toxin proteins that are known to exhibit insecticidal activity. The inventors herein have identified that such smaller toxin molecules, when provided in the diet of a C. elegans nematode, exhibit various levels of inhibitory effects. Surprisingly, it has also been observed that the nematidical activity of these proteins can be imparted through the diet of a cyst nematode more effectively by truncating the proteins to smaller sizes, whether truncated at the C terminus, the N terminus, or both. Truncated versions typically exhibit a mass of from about 14 to about 28-30 kDa, and exhibit improved bioactivity likely because the ability of the cyst nematode to uptake proteins larger than about 30 kDa is limited (Urwin et al. ((1997) Plant J. 12:455) and Bockenhoff & Grundler ((1994) Parasitology 109:249). TIC1501 (about 27 kDa), TIC1503 (about 34 kDa), and TIC1506 (about 36 kDa) represent various fragments of the TIC1201 protein, 1201 being previously shown to exhibit coleopteran toxic effects. Surprisingly, the truncated versions less than 36 kDa exhibit significant nematicidal effects.

[0053] Proteins of the present invention have also been identified de novo, and these include the proteins listed herein as TIC614, TIC615, TIC1277, TIC1278, TIC1308, TIC1310, TIC1311, TIC1324, TIC1407, TIC1408, and TIC1442. Such proteins were identified by various methods, whether being directly amplified from various Bt strain genomes, or identified by high throughput sequence analysis of various Bt genomes. In either case, genomic DNA segments are obtained and analyzed using bioinformatic techniques that result in the identification of all or portions of open reading frames encoding protein segments. The resulting protein segments are then characterized versus all known protein sequences in the art, and to the extent that there is any similarity to a toxin molecule, the complete sequence of the open reading frame encoding the protein is obtained. Proteins that are identified that exhibit a mass of less than about 40 kDa, or preferably less than about 30 kDa are then evaluated in a C. elegans assay to determine if any effects are observed relative to C. elegans survival. Toxins exhibiting nematicidal properties are then evaluated for other pesticidal properties, particularly insecticidal activity. Surprisingly, the above referenced proteins all exhibited nematicidal activity, and some exhibited insecticidal activity as reported in the examples below.

EXAMPLES

[0054] The following examples are illustrative of the invention, which may be embodied in various forms and are not to be interpreted as limiting the scope or content of the disclosure in any way.

Example 1

DNA Molecules Encoding Bt Toxin Proteins

[0055] Toxin ET34 (SEQ ID NO:1) has been previously described (U.S. Pat. No. 6,063,756). A secretion signal from the gene P139 (First 75 nucleotide of SEQ ID NO:1 from the WIPO Publication Number WO9408010) was operably linked to the 5' end of the ET34 (SEQ ID NO:5) to enable its secretion outside the plasma membrane to avoid potential toxicity to the plant cell and to allow easy access of the protein to the pest.

[0056] TIC1506 (SEQ ID NO:9), TIC1501 (SEQ ID NO:13), and TIC 1503 (SEQ ID NO:17) are Bt nucleotide of various fragments of TIC1201 which is 364 amino acids long as set forth in SEQ ID NO 6 of US Patent Application Publication Number US2006-0191034 A1. TIC1506 is 321 amino acids long without the putative N terminal signal peptide of TIC1201 and contains amino acid 44 to 364 of TIC1201 with a methionine residue substituted for the native alanine residue at amino acid position 44. TIC1501 is 227 amino acids long without the putative N terminal signal peptide and a portion of the C terminal of TIC1201 and contains amino acids 44 to 270 of TIC1201 with a methionine residue substituted for the native alanine residue at position 44. TIC1503 is 301 amino acids long without the putative N terminal signal peptide and a portion of the C terminal of TIC1201 and contains amino acids 44 to 344 of TIC1201 with a methionine residue substituted for the native alanine residue at position 44.

[0057] Proteins exhibiting pesticidal properties have been identified in various Bt strains. Open reading frames encoding the amino acid sequences, TIC614 (SEQ ID NO: 22), TIC615 (SEQ ID NO: 26), TIC1277 (SEQ ID NO: 30), TIC1278 (SEQ ID NO: 34), TIC1310 (SEQ ID NO: 38), TIC1311 (SEQ ID NO: 42), TIC1324 (SEQ ID NO: 46), TIC1407 (SEQ ID NO: 50), and TIC1408 (SEQ ID NO: 54), exhibiting various degrees of homology to previously known Bt toxin segments were identified. Complete forward and reverse sequence analysis of such open reading frames resulted in the identification of deduced amino acid compositions that exhibit the size and potential for pesticidal (nematicidal and/or insecticidal) activity.

[0058] Proteins exhibiting pesticidal properties have been identified in various Bt strains. Open reading frames encoding the amino acid sequences, TIC1308 (SEQ ID NO: 58) and TIC1442 (SEQ ID NO: 60), exhibiting various degrees of homology to previously known Bt toxin segments were identified. Complete forward and reverse sequence analysis of such open reading frames resulted in the identification of deduced amino acid compositions that exhibit the size and potential for pesticidal (nematicidal and/or insecticidal) activity.

Example 2

Expression of Pesticidal Polypeptides from Polynucleotides

[0059] Open reading frames of ET34 (SEQ ID NO: 1), P139-ET34 (SEQ ID NO: 5), TIC1506 (SEQ ID NO:9), TIC1501 (SEQ ID NO:13), TIC 1503 (SEQ ID NO:17), TIC614 (SEQ ID NO:21), TIC615 (SEQ ID NO:25), TIC1277 (SEQ ID NO:29), TIC1278 (SEQ ID NO:33), TIC1310 (SEQ ID NO:37), TIC1311 (SEQ ID NO:41), TIC1324 (SEQ ID NO:45), TIC1407 (SEQ ID NO:49), TIC1408 (SEQ ID NO:53), TIC1308 (SEQ ID NO: 57), and TIC1442 (SEQ ID NO: 59) encoding the deduced amino acid compositions exemplified in Example 1 were cloned into a Bt/E. coli shuttle plasmid enabling the expression of the deduced amino acid composition in either an acrystalliferous Bt strain or in an E. coli bacterium. Recombinant plasmids were transformed into an acrystalliferous Bt expression host after confirming the DNA sequence of the polynucleotide encoding the polypeptide. The gene of interest was cloned downstream of either a Bacillus vegetative stage or sporulation stage specific promoter to allow the protein to be expressed respectively during vegetative growth or during sporulation of the recombinant Bt strain. Conditions for vegetative expression of the protein included growing the cells for 24-48 hrs in Terrific Broth medium at 25-28° C. Crystal formation is one characteristic of certain Bt toxin proteins. Proteins that were confirmed to produce crystals when expressed in the acrystalliferous Bt strain were further evaluated. Certain proteins accumulated in the cells and/or were secreted into the culture medium. Both the cell pellets and the culture were analyzed by SDS-PAGE for expression of the expected protein. Conditions for expression from a sporulation specific promoter included growing the cells for 96 hrs at 25-28° C. in C2 medium. Protein crystals were formed during sporulation and released from lysed cells as sporulation was completed. Spores and crystals were collected by centrifugation at 4000×g for 20 minutes, resuspended in wash buffer (10 mM Tris, 0.1 mM EDTA and 0.005% Triton X100 pH 6.8) and collected again by centrifugation. The spore-crystals pellets were then resuspended in 1/10^th of the original culture volume. The 10× concentrated spore-crystal preparation were analyzed by SDS-PAGE the presence of the expected protein.

Example 3

Nematode and Insect Bioassays

[0060] C. elegans feeding screens have been successfully used to identify plant pathogenic nematode-active toxins, for example for SCN and RKN from the order Tylenchida (Wei et al., 2003, PNAS, USA, 100: 2760). The proteins of the present invention were expressed and provided in the diet of a C. elegans nematode, essentially following the method of Wei et al. Efficacy was scored on a scale of 1-3, where a score of 1 represents normal health and reproduction of C. elegans, and a score of 3 represents no reproduction or poor health of C. elegans. Toxins ET34 (SEQ ID NO: 2), TIC1501 (SEQ ID NO: 14), TIC1503 (SEQ ID NO: 18), TIC614 (SEQ ID NO:22), TIC615 (SEQ ID NO:26), TIC1277 (SEQ ID NO:30), TIC1278 (SEQ ID NO:34), TIC1310 (SEQ ID NO:38), TIC1311 (SEQ ID NO:42), TIC1324 (SEQ ID NO:46), and TIC1407 (SEQ ID NO:50), each exhibited a score of 3 where as TIC1408 (SEQ ID NO:54) exhibited a score of 2.75. Proteins TIC1308 (SEQ ID NO: 58) and TIC1442 (SEQ ID NO: 60), each exhibited a score of 1. Proteins were expressed from a vegetative specific promoter and fed to insects by applying sporulated bacterial cells or culture supernatant to artificial insect diet. For polypeptides expressed from a sporulation specific promoter a 10-20× spore-crystal preparation with about 500-4000 ppm protein was applied to the insect diet. Stunting or mortality was observed on one or more of these insects: CEW (Corn Ear Worm); SCR (Southern corn Root worm); WCR (western corn root worm); ECB (European corn borer); WTPB (Western tarnished plant bug); TPB (Tarnished plant bug); FAW (Fall army worm); CPB (Colorado potato beetle). TIC1277 and TIC1311 were found to cause significant stunting of ECB and TIC1310 was found to cause significant stunting of WTPB and significant stunting and mortality of CPB.

Example 4

Transformed Plants

[0061] Nucleotide segments encoding TIC1506 (SEQ ID NO:11), TIC1503 (SEQ ID NO: 19), TIC614 (SEQ ID NO:3), TIC1324 (SEQ ID NO:27), TIC1407 (SEQ ID NO:31), TIC1408 (SEQ ID NO:35), are codon-optimized for plant expression and operably linked to one or more plant functional promoters and introduced into plant cells. Recombinant plants are regenerated from such transformed plant cells, and the regenerated plants are evaluated for resistance to pest infestation, such as insect tolerance and/or plant pathogenic nematode tolerance.

[0062] Nucleotide segments encoding ET34 (SEQ ID NO:3), ET34+P139 secretion signal (SEQ ID NO:7), TIC1501 (SEQ ID NO:15), TIC615 (SEQ ID NO:7), TIC1277 (SEQ ID NO:11), TIC1278 (SEQ ID NO:15), TIC1310 (SEQ ID NO:19), TIC1311 (SEQ ID NO:23), TIC1308 (SEQ ID NO: 61), and TIC1442 (SEQ ID NO: 63) were codon-optimized for plant expression and operably linked to one or more plant functional promoters and introduced into plant cells. Recombinant plants were regenerated from such transformed plant cells, and the regenerated plants were evaluated for resistance to pest infestation, such as insect tolerance and/or plant pathogenic nematode tolerance.

Example 5

Transformation of Soybean

[0063] This example describes a method of producing transgenic soybean plants and transgenic plant parts such as seeds. Other methods are known in the art of plant cell transformation that can be applied to transform plant cells and regenerate transgenic plants using the recombinant constructs of the invention. The methods of obtaining transgenic soybean plants and seeds are used as essentially disclosed in US Patent Application Publication Number US2009-0138985A1. Briefly, Agrobacterium containing a construct of Example 5 are grown in Luria Burtani (LB) media containing spectinomycin at about 28° C. for over night. The bacterial culture is centrifuged, pellet washed, and resuspended in INO medium for inoculating wet or dry mature embryos explants. The explants are mixed with the Agrobacterium cell suspensions and briefly exposed to sonication energy from a standard laboratory water bath cleaning sonicator. The explants are drained of any liquid and transferred to containers containing filter paper moistened with INO media and co-cultured in a lighted chamber at about 16 hours of light (≧5 uE) at about 23° to 28 C.° for 1 to 5 days. After co-culture, the explants are placed directly onto regeneration media containing a selective agent such as spectinomycin from about 7 to about 42 days. The cultures are subsequently transferred to a media suitable for the recovery of transformed plantlets. Spectinomycin resistant shoots that have green buds or leaves are considered transformed and placed in soil or on a soil substitute for rooting in the presence or absence of the selective agent. Progeny transgenic plants and seed are selected that provide pest resistance, especially nematode resistance.

Example 6

Testing of Transgenic Plant for Soybean Cyst Nematode (SCN) Resistance

[0064] An SCN pot assay was used to evaluate the resistance of transgenic soybean plants comprising one or more of the polynucleotide sequences of SEQ ID NOs: 3, 7, and 15 to infection by and reproduction of the SCN (Heterodera glycines) on roots. Three or four inch diameter square pots were filled with clean sand and watered thoroughly. Transgenic and control soybean seeds, or alternatively any rooted plant parts, were planted one per pot in the center of the pot and watered well to remove air pockets. The pots were incubated in the greenhouse or growth chamber at 20° C. to 30° C. until the plants reached a suitable age for inoculation. Soybeans started from seed were typically inoculated 2-3 weeks after planting, while transplants were inoculated 1-3 days after planting. The test inoculum consisted of eggs from ripe H. glycines cysts collected from the soil and roots of infested soybean plants. An 80 micron mesh sieve was used to collect the cysts, which were then crushed in a Tenbroeck glass tissue homogenizer to release the eggs. The eggs were further purified by sieving and centrifugation over 40 percent sucrose solution at 4000 RPM for 5 minutes. Inoculum for an experiment consisted of water containing 500 vermiform eggs per mL. Five mL of the egg suspension was applied over the surface of the sand containing the test plants and the eggs were lightly watered in. The test plants were then returned to the greenhouse or growth chamber and incubated for 3-4 weeks to allow for root infection and cyst formation. The roots were then harvested. The severity of nematode infection was measured by counting the number of nematode cysts adhering to the root system.

[0065] Transgenic soybean plants comprising SEQ ID NO: 3 were tested in six different constructs, where in each construct SEQ ID NO: 3 was operably linked to a different promoter. Transgenic soybean plants comprising SEQ ID NO: 7 were tested in two different constructs, each construct having a different promoter. Transgenic soybean plants comprising SEQ ID NO: 15 were tested expressed from one construct.

[0066] Table 1 reports data illustrating plants from multiple events per multiple constructs that were evaluated for and determined to have significant cyst reduction against SCN when compared to the untransformed soybean cultivar. The number of plant roots tested was about equally distributed among the number of events tested.

TABLE-US-00001 TABLE 1 Severity of Soybean Cyst Nematode cyst infection on soybean plant roots Protein SEQ ID Construct Number of Number of plant Name NO: Name Events tested roots tested Result (compared to control non-transgenic plants) ET34 3 128213 14 177 1 of 14 events showed 35.4% cyst reduction ET34 3 126168 15 389 7 of 15 events showed more than 46.7% cyst reduction ET34 3 126626 15 191 0 of 15 events showed significant cyst reduction ET34 3 126630 16 204 1 of 16 events showed 42.0% cyst reduction ET34 3 127056 15 191 3 of 15 events showed more than 41.5% cyst reduction ET34 3 128296 14 168 2 of 14 events showed more than 33% cyst reduction P139 + ET34 7 126169 15 176 0 of 15 events showed the significant cyst reduction P139 + ET34 7 126628 16 192 0 of 16 events showed the significant cyst reduction TIC1501 15 133535 5 18 3 of 5 events showed more than 45% cyst reduction

Example 7

Testing of Transgenic Arabidopsis Plant for Beet Cyst Nematode (BCN) Resistance

[0067] Transgenic Arabidopsis seeds and plants comprising one or more of the polynucleotide sequences of SEQ ID NOs 3, 15, 27, 31, 35, 39, 43, 61, and 63 were produced by the method of Clough et al., 1998 (Plant J. 16:735-743) and tested for Beet Cyst Nematode (BCN) resistance by the method of Sijmons et al., 1991 (Plant J. 1: 245-254) and Vaghchhipawala et al., 2004 (Genome 47: 404-13).

[0068] Arabidopsis (variety Columbia-0) seeds were surface sterilized and rinsed with sterile water and plated on B5 medium. Plates were incubated at 23-25° C. with a 16 hour light/8 hour dark cycle for 7-10 days. BCN eggs were placed on the sterile filter paper and hatched in 5 mM ZnSO4 solution for 5-7 days at 25° C. J2 stage juvenile nematodes were collected, rinsed in sterile water, and treated with 0.5% chlorhexidine diacetate for 10-15 minutes. Treated juvenile J2 nematodes were collected and rinsed twice in sterile water and stored in sterile water for infestation purposes.

[0069] For the infestation assay, about 10-15 Arabidopsis seeds were sprinkled on steamed sand in a pot and covered with a clear plastic dome. Several such dome/flat combos were placed in a flat and then covered with a black tray and transferred to a cold room for vernalization. On day 4, the flat was taken out of the cold room, the black tray is removed, and the flat was placed in a growth chamber for acclimating seeds at 26° C., 70% humidity, 140-180 μE light, 12 hours day length. The pots were watered and fertilized as needed. Three weeks after planting, the Arabidopsis plants were inoculated with 3,000 BCN eggs. About 35 days after inoculation the plants were harvested and cysts extracted by washing the plant's roots in a bucket of water and filtering the water through a 16 mesh sieve on top of a 50 mesh sieve. The cysts were collected off the top of the 50 mesh sieve and counted. Plants with lesser number of cysts compared to non-transgenic or transgenic control were considered resistant to BCN.

[0070] Table 2 reports data illustrating plants from multiple events per multiple constructs that were evaluated for and determined to have lesser number of BCN cysts compared to the untransformed Arabidopsis parental background. The number of plant roots tested was nearly equally distributed among the number of events tested.

TABLE-US-00002 TABLE 2 Severity of Beet Cyst Nematode cyst infection on Arabidopsis thaliana plant roots Resulting number Number Number of events having a SEQ of of plant lower mean cyst Protein ID Construct Events roots count compared to Name NO: Name tested tested non-transgenic control ET34 3 140057 6 72 3 TIC1501 15 140056 6 72 3 TIC615 27 139822 6 72 4 TIC1277 31 142259 6 71 2 TIC1278 35 141647 6 72 4 TIC1310 39 142255 6 72 1 TIC1311 43 141644 6 71 6 TIC1308 61 141250 6 70 2 TIC1422 63 141205 6 72 2

[0071] Various patent and non-patent publications are cited herein, the disclosures of each of which are incorporated herein by reference in their entireties. Documents cited herein as being available from the World Wide Web at certain internet addresses are also incorporated herein by reference in their entireties.

[0072] As various modifications could be made in the compositions and methods herein described and illustrated without departing from the scope of the invention, it is intended that all matter contained in the foregoing description or shown in the accompanying drawings shall be interpreted as illustrative rather than limiting. Thus, the breadth and scope of the present invention should not be limited by any of the above-described exemplary embodiments, but should be defined only in accordance with the following claims appended hereto and their equivalents.

Sequence CWU 1

641381DNABacillus thuringiensis 1atgacagtat ataacgcaac tttcaccatt aatttctata atgaaggaga atgggggggg 60ccagaaccat atggttatat aaaagcatat cttacaaatc cagatcatga ttttgaaatt 120tggaaacaag atgattgggg gaaaagtact cctgagagaa gtacttatac gcaaacgatt 180aaaataagta gcgacactgg ttcccctata aaccaaatgt gtttttatgg tgatgtgaaa 240gaatacgacg taggaaatgc agatgatatt ctcgcttatc caagtcaaaa agtatgcagt 300acacctggtg taacagtacg acttgatggc gatgagaaag gttcttatgt gacaattaag 360tattccttga ctccagcata a 3812126PRTBacillus thuringiensis 2Met Thr Val Tyr Asn Ala Thr Phe Thr Ile Asn Phe Tyr Asn Glu Gly1 5 10 15Glu Trp Gly Gly Pro Glu Pro Tyr Gly Tyr Ile Lys Ala Tyr Leu Thr 20 25 30Asn Pro Asp His Asp Phe Glu Ile Trp Lys Gln Asp Asp Trp Gly Lys 35 40 45Ser Thr Pro Glu Arg Ser Thr Tyr Thr Gln Thr Ile Lys Ile Ser Ser 50 55 60Asp Thr Gly Ser Pro Ile Asn Gln Met Cys Phe Tyr Gly Asp Val Lys65 70 75 80Glu Tyr Asp Val Gly Asn Ala Asp Asp Ile Leu Ala Tyr Pro Ser Gln 85 90 95Lys Val Cys Ser Thr Pro Gly Val Thr Val Arg Leu Asp Gly Asp Glu 100 105 110Lys Gly Ser Tyr Val Thr Ile Lys Tyr Ser Leu Thr Pro Ala 115 120 1253381DNAArtificial sequencerecombinant polynucleotide encoding an ET34 protein. 3atgactgttt acaacgctac tttcactatt aacttctaca acgagggtga gtggggtgga 60cctgagcctt acggttacat taaggcttac ttgactaatc cagatcatga tttcgagatt 120tggaagcaag atgattgggg caagagcact ccagagcgct ctacctacac ccaaaccatt 180aagatttcat ccgataccgg atcacccatc aatcagatgt gcttctacgg agatgttaag 240gagtatgatg ttggcaatgc tgacgacatc cttgcctatc cctcccagaa ggtctgtagt 300acacccggcg tcacagtgag gctcgacggc gacgagaagg gttcctatgt aaccatcaag 360tattcgctga cgcctgcatg a 3814126PRTArtificial sequencean amino acid sequence translation of SEQ ID NO 3 from nucleotide position 1 through nucleotide position 378 4Met Thr Val Tyr Asn Ala Thr Phe Thr Ile Asn Phe Tyr Asn Glu Gly1 5 10 15Glu Trp Gly Gly Pro Glu Pro Tyr Gly Tyr Ile Lys Ala Tyr Leu Thr 20 25 30Asn Pro Asp His Asp Phe Glu Ile Trp Lys Gln Asp Asp Trp Gly Lys 35 40 45Ser Thr Pro Glu Arg Ser Thr Tyr Thr Gln Thr Ile Lys Ile Ser Ser 50 55 60Asp Thr Gly Ser Pro Ile Asn Gln Met Cys Phe Tyr Gly Asp Val Lys65 70 75 80Glu Tyr Asp Val Gly Asn Ala Asp Asp Ile Leu Ala Tyr Pro Ser Gln 85 90 95Lys Val Cys Ser Thr Pro Gly Val Thr Val Arg Leu Asp Gly Asp Glu 100 105 110Lys Gly Ser Tyr Val Thr Ile Lys Tyr Ser Leu Thr Pro Ala 115 120 1255453DNAArtificial sequencerecombinant polynucleotide encoding a P139 secretion signal peptide (nucleotide position 1-75) fused in frame to a native Bt nucleotide sequence encoding a ET34 protein (nucleotide position 76-450) 5atgaggagtt tcgccgtctt gttgccgctt ctcgtcacct tctgcgtggt ggcgcctcct 60tccgacgccg ccaccacagt atataacgca actttcacca ttaatttcta taatgaagga 120gaatgggggg ggccagaacc atatggttat ataaaagcat atcttacaaa tccagatcat 180gattttgaaa tttggaaaca agatgattgg gggaaaagta ctcctgagag aagtacttat 240acgcaaacga ttaaaataag tagcgacact ggttccccta taaaccaaat gtgtttttat 300ggtgatgtga aagaatacga cgtaggaaat gcagatgata ttctcgctta tccaagtcaa 360aaagtatgca gtacacctgg tgtaacagta cgacttgatg gcgatgagaa aggttcttat 420gtgacaatta agtattcctt gactccagca taa 4536150PRTArtificial sequencean amino acid sequence translation of SEQ ID NO 5. 6Met Arg Ser Phe Ala Val Leu Leu Pro Leu Leu Val Thr Phe Cys Val1 5 10 15Val Ala Pro Pro Ser Asp Ala Ala Thr Thr Val Tyr Asn Ala Thr Phe 20 25 30Thr Ile Asn Phe Tyr Asn Glu Gly Glu Trp Gly Gly Pro Glu Pro Tyr 35 40 45Gly Tyr Ile Lys Ala Tyr Leu Thr Asn Pro Asp His Asp Phe Glu Ile 50 55 60Trp Lys Gln Asp Asp Trp Gly Lys Ser Thr Pro Glu Arg Ser Thr Tyr65 70 75 80Thr Gln Thr Ile Lys Ile Ser Ser Asp Thr Gly Ser Pro Ile Asn Gln 85 90 95Met Cys Phe Tyr Gly Asp Val Lys Glu Tyr Asp Val Gly Asn Ala Asp 100 105 110Asp Ile Leu Ala Tyr Pro Ser Gln Lys Val Cys Ser Thr Pro Gly Val 115 120 125Thr Val Arg Leu Asp Gly Asp Glu Lys Gly Ser Tyr Val Thr Ile Lys 130 135 140Tyr Ser Leu Thr Pro Ala145 1507453DNAArtificial sequencea recombinant polynucleotide sequence encoding a P139 secretion signal peptide (nucleotide position 1-75) fused in frame to a synthetic nucleotide sequence encoding a ET34 protein (nucleotide position 76-450) 7atgaggagtt tcgccgtctt gttgccgctt ctcgtcacct tctgcgtggt ggcgcctcct 60tccgacgccg ccaccactgt ttacaacgct actttcacta ttaacttcta caacgagggt 120gagtggggtg gacctgagcc ttacggttac attaaggctt acttgactaa tccagatcat 180gatttcgaga tttggaagca agatgattgg ggcaagagca ctccagagcg ctctacctac 240acccaaacca ttaagatttc atccgatacc ggatcaccca tcaatcagat gtgcttctac 300ggagatgtta aggagtatga tgttggcaat gctgacgaca tccttgccta tccctcccag 360aaggtctgta gtacacccgg cgtcacagtg aggctcgacg gcgacgagaa gggttcctat 420gtaaccatca agtattcgct gacgcctgca tga 4538150PRTArtificial sequencean amino acid sequence translation of SEQ ID NO 7. 8Met Arg Ser Phe Ala Val Leu Leu Pro Leu Leu Val Thr Phe Cys Val1 5 10 15Val Ala Pro Pro Ser Asp Ala Ala Thr Thr Val Tyr Asn Ala Thr Phe 20 25 30Thr Ile Asn Phe Tyr Asn Glu Gly Glu Trp Gly Gly Pro Glu Pro Tyr 35 40 45Gly Tyr Ile Lys Ala Tyr Leu Thr Asn Pro Asp His Asp Phe Glu Ile 50 55 60Trp Lys Gln Asp Asp Trp Gly Lys Ser Thr Pro Glu Arg Ser Thr Tyr65 70 75 80Thr Gln Thr Ile Lys Ile Ser Ser Asp Thr Gly Ser Pro Ile Asn Gln 85 90 95Met Cys Phe Tyr Gly Asp Val Lys Glu Tyr Asp Val Gly Asn Ala Asp 100 105 110Asp Ile Leu Ala Tyr Pro Ser Gln Lys Val Cys Ser Thr Pro Gly Val 115 120 125Thr Val Arg Leu Asp Gly Asp Glu Lys Gly Ser Tyr Val Thr Ile Lys 130 135 140Tyr Ser Leu Thr Pro Ala145 1509966DNABacillus thuringiensis 9gcaataactc catatgctga atcttatatt gatactgttc aagatagaat gaaacaaaga 60gatagggaat caaaactaac tggtaaacca ataaatatgc aagaacaaat aatagatgga 120tggtttttag ctagattctg gatatttaaa gatcaaaata acaatcatca aacaaataga 180tttatatcct ggtttaaaga taatcttgct agttcgaagg ggtatgacag tatagcagaa 240caaatgggct taaaaataga agcattaaat gatatggatg taacaaatat tgattataca 300tctaaaacag gtgataccat atataatgga atttctgaac taacaaatta tacaggaaca 360acccaaaaaa tgaaaaccga tagttttcaa agagattata caaaatctga atccacttca 420gtaacaaatg ggttacaatt aggatttaaa gttgctgcta agggagtagt tgcattagca 480ggtgcagatt ttgaaacaag tgttacctat aatttatcat ctactacaac tgaaacaaat 540acaatatcgg ataagtttac tgttccatct caagaagtta cattatcccc aggacataaa 600gcagtggtga aacatgattt gagaaaaatg gtgtattttg ggactcatga tttaaagggt 660gatttaaaag taggttttaa tgataaagag attgtacaaa aatttattta tccaaattat 720agatcaattg atttatctga tattcgtaaa acaatgattg aaattgataa atggaatcat 780gtaaatacca ttgactttta tcaattagtt ggagttaaaa atcatataaa aaatggtgat 840actttatata tagatacccc ggccgaattt acatttaatg gagctaatcc atattataga 900gcaacattta cagaatacga cgagaacgga aatcctgttc aaacaaagat tttaagtgga 960aattaa 96610321PRTArtificial sequencean amino acid sequence translation of SEQ ID NO 9. 10Met Ile Thr Pro Tyr Ala Glu Ser Tyr Ile Asp Thr Val Gln Asp Arg1 5 10 15Met Lys Gln Arg Asp Arg Glu Ser Lys Leu Thr Gly Lys Pro Ile Asn 20 25 30Met Gln Glu Gln Ile Ile Asp Gly Trp Phe Leu Ala Arg Phe Trp Ile 35 40 45Phe Lys Asp Gln Asn Asn Asn His Gln Thr Asn Arg Phe Ile Ser Trp 50 55 60Phe Lys Asp Asn Leu Ala Ser Ser Lys Gly Tyr Asp Ser Ile Ala Glu65 70 75 80Gln Met Gly Leu Lys Ile Glu Ala Leu Asn Asp Met Asp Val Thr Asn 85 90 95Ile Asp Tyr Thr Ser Lys Thr Gly Asp Thr Ile Tyr Asn Gly Ile Ser 100 105 110Glu Leu Thr Asn Tyr Thr Gly Thr Thr Gln Lys Met Lys Thr Asp Ser 115 120 125Phe Gln Arg Asp Tyr Thr Lys Ser Glu Ser Thr Ser Val Thr Asn Gly 130 135 140Leu Gln Leu Gly Phe Lys Val Ala Ala Lys Gly Val Val Ala Leu Ala145 150 155 160Gly Ala Asp Phe Glu Thr Ser Val Thr Tyr Asn Leu Ser Ser Thr Thr 165 170 175Thr Glu Thr Asn Thr Ile Ser Asp Lys Phe Thr Val Pro Ser Gln Glu 180 185 190Val Thr Leu Ser Pro Gly His Lys Ala Val Val Lys His Asp Leu Arg 195 200 205Lys Met Val Tyr Phe Gly Thr His Asp Leu Lys Gly Asp Leu Lys Val 210 215 220Gly Phe Asn Asp Lys Glu Ile Val Gln Lys Phe Ile Tyr Pro Asn Tyr225 230 235 240Arg Ser Ile Asp Leu Ser Asp Ile Arg Lys Thr Met Ile Glu Ile Asp 245 250 255Lys Trp Asn His Val Asn Thr Ile Asp Phe Tyr Gln Leu Val Gly Val 260 265 270Lys Asn His Ile Lys Asn Gly Asp Thr Leu Tyr Ile Asp Thr Pro Ala 275 280 285Glu Phe Thr Phe Asn Gly Ala Asn Pro Tyr Tyr Arg Ala Thr Phe Thr 290 295 300Glu Tyr Asp Glu Asn Gly Asn Pro Val Gln Thr Lys Ile Leu Ser Gly305 310 315 320Asn11966DNAArtificial sequencerecombinant polynucleotide encoding a TIC1506 protein. 11atgattactc cttacgctga gtcttacatt gatactgttc aagatcgcat gaagcaacgt 60gatcgtgagt ctaagttgac tggcaagcct attaacatgc aagagcaaat tatcgacggt 120tggttcctag ccagattctg gatcttcaag gatcagaaca ataaccacca gaccaaccgc 180ttcatcagtt ggttcaagga taacttggct agttctaagg gttacgatag tattgctgag 240caaatgggtt tgaagattga ggctttgaac gatatggatg ttaccaacat tgattacact 300tccaagactg gtgatacgat ctacaacggt atttctgagt tgactaacta cactggtact 360actcagaaga tgaagaccga cagcttccag agggattaca ccaagtctga gtcaacctca 420gttaccaacg gacttcagct tggattcaag gttgcagcca agggagttgt ggcacttgct 480ggagcagatt tcgagacctc agttacctac aacctttcat ccaccacaac cgaaacgaac 540accatctccg ataagtttac cgttccatcc caggaagtga cactttcccc aggacacaag 600gccgtggtca agcacgatct caggaagatg gtgtacttcg gaacacacga cctcaaaggc 660gacctcaaag tgggctttaa cgacaaagaa atcgtgcaga agttcatcta tccaaattat 720cgcagcatcg acctcagcga catccgaaag acaatgatcg aaatcgacaa atggaatcac 780gtcaacacaa tcgacttcta tcaactggtc ggcgtaaaga accacatcaa gaacggcgac 840acactctaca tcgacacacc cgccgagttt acattcaatg gggccaatcc ctattatcgg 900gcgacattca cggaatacga cgagaatggg aatccggtac agacgaagat cctgtcgggg 960aattga 96612321PRTArtificial sequencean amino acid sequence translation of SEQ ID NO 11. 12Met Ile Thr Pro Tyr Ala Glu Ser Tyr Ile Asp Thr Val Gln Asp Arg1 5 10 15Met Lys Gln Arg Asp Arg Glu Ser Lys Leu Thr Gly Lys Pro Ile Asn 20 25 30Met Gln Glu Gln Ile Ile Asp Gly Trp Phe Leu Ala Arg Phe Trp Ile 35 40 45Phe Lys Asp Gln Asn Asn Asn His Gln Thr Asn Arg Phe Ile Ser Trp 50 55 60Phe Lys Asp Asn Leu Ala Ser Ser Lys Gly Tyr Asp Ser Ile Ala Glu65 70 75 80Gln Met Gly Leu Lys Ile Glu Ala Leu Asn Asp Met Asp Val Thr Asn 85 90 95Ile Asp Tyr Thr Ser Lys Thr Gly Asp Thr Ile Tyr Asn Gly Ile Ser 100 105 110Glu Leu Thr Asn Tyr Thr Gly Thr Thr Gln Lys Met Lys Thr Asp Ser 115 120 125Phe Gln Arg Asp Tyr Thr Lys Ser Glu Ser Thr Ser Val Thr Asn Gly 130 135 140Leu Gln Leu Gly Phe Lys Val Ala Ala Lys Gly Val Val Ala Leu Ala145 150 155 160Gly Ala Asp Phe Glu Thr Ser Val Thr Tyr Asn Leu Ser Ser Thr Thr 165 170 175Thr Glu Thr Asn Thr Ile Ser Asp Lys Phe Thr Val Pro Ser Gln Glu 180 185 190Val Thr Leu Ser Pro Gly His Lys Ala Val Val Lys His Asp Leu Arg 195 200 205Lys Met Val Tyr Phe Gly Thr His Asp Leu Lys Gly Asp Leu Lys Val 210 215 220Gly Phe Asn Asp Lys Glu Ile Val Gln Lys Phe Ile Tyr Pro Asn Tyr225 230 235 240Arg Ser Ile Asp Leu Ser Asp Ile Arg Lys Thr Met Ile Glu Ile Asp 245 250 255Lys Trp Asn His Val Asn Thr Ile Asp Phe Tyr Gln Leu Val Gly Val 260 265 270Lys Asn His Ile Lys Asn Gly Asp Thr Leu Tyr Ile Asp Thr Pro Ala 275 280 285Glu Phe Thr Phe Asn Gly Ala Asn Pro Tyr Tyr Arg Ala Thr Phe Thr 290 295 300Glu Tyr Asp Glu Asn Gly Asn Pro Val Gln Thr Lys Ile Leu Ser Gly305 310 315 320Asn13681DNABacillus thuringiensis 13gcaataactc catatgctga atcttatatt gatactgttc aagatagaat gaaacaaaga 60gatagggaat caaaactaac tggtaaacca ataaatatgc aagaacaaat aatagatgga 120tggtttttag ctagattctg gatatttaaa gatcaaaata acaatcatca aacaaataga 180tttatatcct ggtttaaaga taatcttgct agttcgaagg ggtatgacag tatagcagaa 240caaatgggct taaaaataga agcattaaat gatatggatg taacaaatat tgattataca 300tctaaaacag gtgataccat atataatgga atttctgaac taacaaatta tacaggaaca 360acccaaaaaa tgaaaaccga tagttttcaa agagattata caaaatctga atccacttca 420gtaacaaatg ggttacaatt aggatttaaa gttgctgcta agggagtagt tgcattagca 480ggtgcagatt ttgaaacaag tgttacctat aatttatcat ctactacaac tgaaacaaat 540acaatatcgg ataagtttac tgttccatct caagaagtta cattatcccc aggacataaa 600gcagtggtga aacatgattt gagaaaaatg gtgtattttg ggactcatga tttaaagggt 660gatttaaaag taggttttaa t 68114227PRTArtificial sequencean amino acid sequence translation of SEQ ID NO 13. 14Met Ile Thr Pro Tyr Ala Glu Ser Tyr Ile Asp Thr Val Gln Asp Arg1 5 10 15Met Lys Gln Arg Asp Arg Glu Ser Lys Leu Thr Gly Lys Pro Ile Asn 20 25 30Met Gln Glu Gln Ile Ile Asp Gly Trp Phe Leu Ala Arg Phe Trp Ile 35 40 45Phe Lys Asp Gln Asn Asn Asn His Gln Thr Asn Arg Phe Ile Ser Trp 50 55 60Phe Lys Asp Asn Leu Ala Ser Ser Lys Gly Tyr Asp Ser Ile Ala Glu65 70 75 80Gln Met Gly Leu Lys Ile Glu Ala Leu Asn Asp Met Asp Val Thr Asn 85 90 95Ile Asp Tyr Thr Ser Lys Thr Gly Asp Thr Ile Tyr Asn Gly Ile Ser 100 105 110Glu Leu Thr Asn Tyr Thr Gly Thr Thr Gln Lys Met Lys Thr Asp Ser 115 120 125Phe Gln Arg Asp Tyr Thr Lys Ser Glu Ser Thr Ser Val Thr Asn Gly 130 135 140Leu Gln Leu Gly Phe Lys Val Ala Ala Lys Gly Val Val Ala Leu Ala145 150 155 160Gly Ala Asp Phe Glu Thr Ser Val Thr Tyr Asn Leu Ser Ser Thr Thr 165 170 175Thr Glu Thr Asn Thr Ile Ser Asp Lys Phe Thr Val Pro Ser Gln Glu 180 185 190Val Thr Leu Ser Pro Gly His Lys Ala Val Val Lys His Asp Leu Arg 195 200 205Lys Met Val Tyr Phe Gly Thr His Asp Leu Lys Gly Asp Leu Lys Val 210 215 220Gly Phe Asn22515684DNAArtificial sequencea recombinant polynucleotide sequence encoding a TIC1501 protein. 15atgattactc cttacgctga gtcttacatt gatactgttc aagatcgcat gaagcaacgt 60gatcgtgagt ctaagttgac tggcaagcct attaacatgc aagagcaaat tatcgacggt 120tggttcctag ccagattctg gatcttcaag gatcagaaca ataaccacca gaccaaccgc 180ttcatcagtt ggttcaagga taacttggct agttctaagg gttacgatag tattgctgag 240caaatgggtt tgaagattga ggctttgaac gatatggatg ttaccaacat tgattacact 300tccaagactg gtgatacgat ctacaacggt atttctgagt tgactaacta cactggtact 360actcagaaga tgaagaccga cagcttccag agggattaca ccaagtctga gtcaacctca 420gttaccaacg gacttcagct tggattcaag gttgcagcca agggagttgt ggcacttgct 480ggagcagatt tcgagacctc agttacctac aacctttcat ccaccacaac cgaaacgaac 540accatctccg ataagtttac cgttccatcc caggaagtga cactttcccc aggacacaag

600gccgtggtca agcacgatct caggaagatg gtgtacttcg gaacacacga cctcaaaggc 660gacctcaaag tgggctttaa ctga 68416227PRTArtificial sequencean amino acid sequence translation of SEQ ID NO 15. 16Met Ile Thr Pro Tyr Ala Glu Ser Tyr Ile Asp Thr Val Gln Asp Arg1 5 10 15Met Lys Gln Arg Asp Arg Glu Ser Lys Leu Thr Gly Lys Pro Ile Asn 20 25 30Met Gln Glu Gln Ile Ile Asp Gly Trp Phe Leu Ala Arg Phe Trp Ile 35 40 45Phe Lys Asp Gln Asn Asn Asn His Gln Thr Asn Arg Phe Ile Ser Trp 50 55 60 Phe Lys Asp Asn Leu Ala Ser Ser Lys Gly Tyr Asp Ser Ile Ala Glu65 70 75 80Gln Met Gly Leu Lys Ile Glu Ala Leu Asn Asp Met Asp Val Thr Asn 85 90 95Ile Asp Tyr Thr Ser Lys Thr Gly Asp Thr Ile Tyr Asn Gly Ile Ser 100 105 110Glu Leu Thr Asn Tyr Thr Gly Thr Thr Gln Lys Met Lys Thr Asp Ser 115 120 125Phe Gln Arg Asp Tyr Thr Lys Ser Glu Ser Thr Ser Val Thr Asn Gly 130 135 140Leu Gln Leu Gly Phe Lys Val Ala Ala Lys Gly Val Val Ala Leu Ala145 150 155 160Gly Ala Asp Phe Glu Thr Ser Val Thr Tyr Asn Leu Ser Ser Thr Thr 165 170 175Thr Glu Thr Asn Thr Ile Ser Asp Lys Phe Thr Val Pro Ser Gln Glu 180 185 190Val Thr Leu Ser Pro Gly His Lys Ala Val Val Lys His Asp Leu Arg 195 200 205Lys Met Val Tyr Phe Gly Thr His Asp Leu Lys Gly Asp Leu Lys Val 210 215 220Gly Phe Asn22517903DNABacillus thuringiensis 17gcaataactc catatgctga atcttatatt gatactgttc aagatagaat gaaacaaaga 60gatagggaat caaaactaac tggtaaacca ataaatatgc aagaacaaat aatagatgga 120tggtttttag ctagattctg gatatttaaa gatcaaaata acaatcatca aacaaataga 180tttatatcct ggtttaaaga taatcttgct agttcgaagg ggtatgacag tatagcagaa 240caaatgggct taaaaataga agcattaaat gatatggatg taacaaatat tgattataca 300tctaaaacag gtgataccat atataatgga atttctgaac taacaaatta tacaggaaca 360acccaaaaaa tgaaaaccga tagttttcaa agagattata caaaatctga atccacttca 420gtaacaaatg ggttacaatt aggatttaaa gttgctgcta agggagtagt tgcattagca 480ggtgcagatt ttgaaacaag tgttacctat aatttatcat ctactacaac tgaaacaaat 540acaatatcgg ataagtttac tgttccatct caagaagtta cattatcccc aggacataaa 600gcagtggtga aacatgattt gagaaaaatg gtgtattttg ggactcatga tttaaagggt 660gatttaaaag taggttttaa tgataaagag attgtacaaa aatttattta tccaaattat 720agatcaattg atttatctga tattcgtaaa acaatgattg aaattgataa atggaatcat 780gtaaatacca ttgactttta tcaattagtt ggagttaaaa atcatataaa aaatggtgat 840actttatata tagatacccc ggccgaattt acatttaatg gagctaatcc atattataga 900gca 90318301PRTArtificial sequencean amino acid sequence translation of SEQ ID NO 17. 18Met Ile Thr Pro Tyr Ala Glu Ser Tyr Ile Asp Thr Val Gln Asp Arg1 5 10 15Met Lys Gln Arg Asp Arg Glu Ser Lys Leu Thr Gly Lys Pro Ile Asn 20 25 30Met Gln Glu Gln Ile Ile Asp Gly Trp Phe Leu Ala Arg Phe Trp Ile 35 40 45Phe Lys Asp Gln Asn Asn Asn His Gln Thr Asn Arg Phe Ile Ser Trp 50 55 60Phe Lys Asp Asn Leu Ala Ser Ser Lys Gly Tyr Asp Ser Ile Ala Glu65 70 75 80Gln Met Gly Leu Lys Ile Glu Ala Leu Asn Asp Met Asp Val Thr Asn 85 90 95Ile Asp Tyr Thr Ser Lys Thr Gly Asp Thr Ile Tyr Asn Gly Ile Ser 100 105 110Glu Leu Thr Asn Tyr Thr Gly Thr Thr Gln Lys Met Lys Thr Asp Ser 115 120 125Phe Gln Arg Asp Tyr Thr Lys Ser Glu Ser Thr Ser Val Thr Asn Gly 130 135 140Leu Gln Leu Gly Phe Lys Val Ala Ala Lys Gly Val Val Ala Leu Ala145 150 155 160Gly Ala Asp Phe Glu Thr Ser Val Thr Tyr Asn Leu Ser Ser Thr Thr 165 170 175Thr Glu Thr Asn Thr Ile Ser Asp Lys Phe Thr Val Pro Ser Gln Glu 180 185 190Val Thr Leu Ser Pro Gly His Lys Ala Val Val Lys His Asp Leu Arg 195 200 205Lys Met Val Tyr Phe Gly Thr His Asp Leu Lys Gly Asp Leu Lys Val 210 215 220Gly Phe Asn Asp Lys Glu Ile Val Gln Lys Phe Ile Tyr Pro Asn Tyr225 230 235 240Arg Ser Ile Asp Leu Ser Asp Ile Arg Lys Thr Met Ile Glu Ile Asp 245 250 255Lys Trp Asn His Val Asn Thr Ile Asp Phe Tyr Gln Leu Val Gly Val 260 265 270Lys Asn His Ile Lys Asn Gly Asp Thr Leu Tyr Ile Asp Thr Pro Ala 275 280 285Glu Phe Thr Phe Asn Gly Ala Asn Pro Tyr Tyr Arg Ala 290 295 30019906DNAArtificial sequencea recombinant polynucleotide sequence encoding a TIC1503 protein. 19atgattactc cttacgctga gtcttacatt gatactgttc aagatcgcat gaagcaacgt 60gatcgtgagt ctaagttgac tggcaagcct attaacatgc aagagcaaat tatcgacggt 120tggttcctag ccagattctg gatcttcaag gatcagaaca ataaccacca gaccaaccgc 180ttcatcagtt ggttcaagga taacttggct agttctaagg gttacgatag tattgctgag 240caaatgggtt tgaagattga ggctttgaac gatatggatg ttaccaacat tgattacact 300tccaagactg gtgatacgat ctacaacggt atttctgagt tgactaacta cactggtact 360actcagaaga tgaagaccga cagcttccag agggattaca ccaagtctga gtcaacctca 420gttaccaacg gacttcagct tggattcaag gttgcagcca agggagttgt ggcacttgct 480ggagcagatt tcgagacctc agttacctac aacctttcat ccaccacaac cgaaacgaac 540accatctccg ataagtttac cgttccatcc caggaagtga cactttcccc aggacacaag 600gccgtggtca agcacgatct caggaagatg gtgtacttcg gaacacacga cctcaaaggc 660gacctcaaag tgggctttaa cgacaaagaa atcgtgcaga agttcatcta tccaaattat 720cgcagcatcg acctcagcga catccgaaag acaatgatcg aaatcgacaa atggaatcac 780gtcaacacaa tcgacttcta tcaactggtc ggcgtaaaga accacatcaa gaacggcgac 840acactctaca tcgacacacc cgccgagttt acattcaatg gggccaatcc ctattatcgg 900gcgtga 90620301PRTArtificial sequencean amino acid sequence translation of SEQ ID NO 19. 20Met Ile Thr Pro Tyr Ala Glu Ser Tyr Ile Asp Thr Val Gln Asp Arg1 5 10 15Met Lys Gln Arg Asp Arg Glu Ser Lys Leu Thr Gly Lys Pro Ile Asn 20 25 30Met Gln Glu Gln Ile Ile Asp Gly Trp Phe Leu Ala Arg Phe Trp Ile 35 40 45Phe Lys Asp Gln Asn Asn Asn His Gln Thr Asn Arg Phe Ile Ser Trp 50 55 60Phe Lys Asp Asn Leu Ala Ser Ser Lys Gly Tyr Asp Ser Ile Ala Glu65 70 75 80Gln Met Gly Leu Lys Ile Glu Ala Leu Asn Asp Met Asp Val Thr Asn 85 90 95Ile Asp Tyr Thr Ser Lys Thr Gly Asp Thr Ile Tyr Asn Gly Ile Ser 100 105 110Glu Leu Thr Asn Tyr Thr Gly Thr Thr Gln Lys Met Lys Thr Asp Ser 115 120 125Phe Gln Arg Asp Tyr Thr Lys Ser Glu Ser Thr Ser Val Thr Asn Gly 130 135 140Leu Gln Leu Gly Phe Lys Val Ala Ala Lys Gly Val Val Ala Leu Ala145 150 155 160Gly Ala Asp Phe Glu Thr Ser Val Thr Tyr Asn Leu Ser Ser Thr Thr 165 170 175Thr Glu Thr Asn Thr Ile Ser Asp Lys Phe Thr Val Pro Ser Gln Glu 180 185 190Val Thr Leu Ser Pro Gly His Lys Ala Val Val Lys His Asp Leu Arg 195 200 205Lys Met Val Tyr Phe Gly Thr His Asp Leu Lys Gly Asp Leu Lys Val 210 215 220Gly Phe Asn Asp Lys Glu Ile Val Gln Lys Phe Ile Tyr Pro Asn Tyr225 230 235 240Arg Ser Ile Asp Leu Ser Asp Ile Arg Lys Thr Met Ile Glu Ile Asp 245 250 255Lys Trp Asn His Val Asn Thr Ile Asp Phe Tyr Gln Leu Val Gly Val 260 265 270Lys Asn His Ile Lys Asn Gly Asp Thr Leu Tyr Ile Asp Thr Pro Ala 275 280 285Glu Phe Thr Phe Asn Gly Ala Asn Pro Tyr Tyr Arg Ala 290 295 30021774DNABacillus thuringiensis 21atggacattc aagatgtaat agaattaatt agtgactatg ccaacaagtg gaaggaacaa 60caaaatggaa gtcgtagtga catgaaattc attgatgcac gctttgcgaa caatgcatta 120aaagtagaag ctgatcgtac tatgtacctg gagcctacag agcaagatat gccagatgta 180aaattagata cccaagtgtt taccaatact tccacacagc ccaaaactgt ccagtttgat 240atgtttggag aatacgatac ttggcaaaca tggagaatcg aagatggtgc aacagaagca 300ggaaaatgtc gctttgccat tgagccattg tttagatcag aggatgtttc tagtccacta 360tctttatttt taaacgacac acttaccctc tcagggaaaa agccattttc cgttgagggt 420cgtgatttta cagttcgacc acgtttcaaa gtaacgggta cactcatggc aaaacccaaa 480acatcaaccc gtgcttttga cgtaaaacgt gacgtcacag gatatgtggg acttgtcaca 540actttagcat ccggagaact gcaagaatcg ttacacaatg tgggcgctat tttccaacaa 600tattatagtc cctatatcga agtaaatggt ggaagagtga catttcatga tcgtggtgaa 660tatcaatcat tagatgtaag cagcatctat attcatattt ttgcagagag cttagatatt 720cccggattaa cggaggaata taatatttat gacctaagca atggacgggt ataa 77422257PRTBacillus thuringiensis 22Met Asp Ile Gln Asp Val Ile Glu Leu Ile Ser Asp Tyr Ala Asn Lys1 5 10 15Trp Lys Glu Gln Gln Asn Gly Ser Arg Ser Asp Met Lys Phe Ile Asp 20 25 30Ala Arg Phe Ala Asn Asn Ala Leu Lys Val Glu Ala Asp Arg Thr Met 35 40 45Tyr Leu Glu Pro Thr Glu Gln Asp Met Pro Asp Val Lys Leu Asp Thr 50 55 60Gln Val Phe Thr Asn Thr Ser Thr Gln Pro Lys Thr Val Gln Phe Asp65 70 75 80Met Phe Gly Glu Tyr Asp Thr Trp Gln Thr Trp Arg Ile Glu Asp Gly 85 90 95Ala Thr Glu Ala Gly Lys Cys Arg Phe Ala Ile Glu Pro Leu Phe Arg 100 105 110Ser Glu Asp Val Ser Ser Pro Leu Ser Leu Phe Leu Asn Asp Thr Leu 115 120 125Thr Leu Ser Gly Lys Lys Pro Phe Ser Val Glu Gly Arg Asp Phe Thr 130 135 140Val Arg Pro Arg Phe Lys Val Thr Gly Thr Leu Met Ala Lys Pro Lys145 150 155 160Thr Ser Thr Arg Ala Phe Asp Val Lys Arg Asp Val Thr Gly Tyr Val 165 170 175Gly Leu Val Thr Thr Leu Ala Ser Gly Glu Leu Gln Glu Ser Leu His 180 185 190Asn Val Gly Ala Ile Phe Gln Gln Tyr Tyr Ser Pro Tyr Ile Glu Val 195 200 205Asn Gly Gly Arg Val Thr Phe His Asp Arg Gly Glu Tyr Gln Ser Leu 210 215 220Asp Val Ser Ser Ile Tyr Ile His Ile Phe Ala Glu Ser Leu Asp Ile225 230 235 240Pro Gly Leu Thr Glu Glu Tyr Asn Ile Tyr Asp Leu Ser Asn Gly Arg 245 250 255Val23774DNAArtificial sequenceSynthetic 23atggacattc aagatgtgat tgagttgatt tctgattacg ccaacaagtg gaaggagcaa 60cagaacggta gtcgtagtga catgaagttc attgatgctc gcttcgccaa caacgctttg 120aaggttgagg ctgatcgcac tatgtaccta gagcctactg agcaagacat gcctgatgtt 180aagttggata ctcaagtgtt cacgaacacc agtacccaac ctaagactgt tcagttcgac 240atgttcggtg agtacgatac ctggcagacc tggagaattg aggatggtgc taccgaggcg 300ggcaagtgca gattcgcaat tgagccattg ttccgctctg aggatgtttc ttcaccactt 360tctctcttcc ttaatgacac ccttaccctt tctggtaaga aaccattcag cgttgaagga 420agggacttta ccgtgaggcc aaggtttaaa gtgaccggaa cactcatggc caaacccaag 480acttcaaccc gtgcctttga cgtgaaacgc gacgtgacag gatacgtggg actcgtgaca 540acgctcgcct caggagaact ccaggaatca ctgcacaatg tcggtgccat ctttcagcag 600tactacagcc cctacatcga agtcaatggc gggcgcgtca cattccatga ccgaggcgag 660tatcagtccc tagacgtatc cagcatctac atccacatct ttgcggaaag cctcgacata 720cctgggctca cggaggagta caacatctac gacctgtcga atgggcgggt atga 77424257PRTArtificial sequenceSynthetic 24Met Asp Ile Gln Asp Val Ile Glu Leu Ile Ser Asp Tyr Ala Asn Lys1 5 10 15Trp Lys Glu Gln Gln Asn Gly Ser Arg Ser Asp Met Lys Phe Ile Asp 20 25 30Ala Arg Phe Ala Asn Asn Ala Leu Lys Val Glu Ala Asp Arg Thr Met 35 40 45Tyr Leu Glu Pro Thr Glu Gln Asp Met Pro Asp Val Lys Leu Asp Thr 50 55 60Gln Val Phe Thr Asn Thr Ser Thr Gln Pro Lys Thr Val Gln Phe Asp65 70 75 80Met Phe Gly Glu Tyr Asp Thr Trp Gln Thr Trp Arg Ile Glu Asp Gly 85 90 95Ala Thr Glu Ala Gly Lys Cys Arg Phe Ala Ile Glu Pro Leu Phe Arg 100 105 110Ser Glu Asp Val Ser Ser Pro Leu Ser Leu Phe Leu Asn Asp Thr Leu 115 120 125Thr Leu Ser Gly Lys Lys Pro Phe Ser Val Glu Gly Arg Asp Phe Thr 130 135 140Val Arg Pro Arg Phe Lys Val Thr Gly Thr Leu Met Ala Lys Pro Lys145 150 155 160Thr Ser Thr Arg Ala Phe Asp Val Lys Arg Asp Val Thr Gly Tyr Val 165 170 175Gly Leu Val Thr Thr Leu Ala Ser Gly Glu Leu Gln Glu Ser Leu His 180 185 190Asn Val Gly Ala Ile Phe Gln Gln Tyr Tyr Ser Pro Tyr Ile Glu Val 195 200 205Asn Gly Gly Arg Val Thr Phe His Asp Arg Gly Glu Tyr Gln Ser Leu 210 215 220Asp Val Ser Ser Ile Tyr Ile His Ile Phe Ala Glu Ser Leu Asp Ile225 230 235 240Pro Gly Leu Thr Glu Glu Tyr Asn Ile Tyr Asp Leu Ser Asn Gly Arg 245 250 255Val 25900DNABacillus thuringiensis 25atggctattt ttaattttga cgcaaaagta gtagagttta ttaattggtg gacagcagag 60ttcggtggca aagatcccag aaatattcaa attggttatg agaatcgaga catcaatgtt 120gttccatcat caagtaaccc tagtgtcaat gtaataccta agttagcaag atcatctgtt 180caagaattgc aaaataatac aagtgtaacg cagacacaag agctggcatt ttcagaaact 240acaactgaaa gtcagtcttc tacaacaaca catggggctt cattttctac aacggtcacc 300tctgttacgc aatttacagc tgaagttaat ttcaaagcga ttggttcctc aattgagcaa 360actatcggcg tttctatgac aggagattat aattacagtt cttcactaac aaaaacaaca 420gaaaagagta gatcttggac tctcacacaa ccggtagttg tcccaccttt ttcacgtgta 480acttgcacat tattgatata taatgcacca ttttcagtac ctgtggactt aaattgtaat 540gtattcggta cacttggcgg agatttttta gctagctaca cttacactgt tattagtact 600ggtcgaacag taaatactag tataactgct agtcaaatga ctcttacctc ttggccaggt 660aaaccatctg aaattattgg tatcgcacca aagcatggac ttatttttaa agggacaggt 720acacaggctg cagtacatgg attatactca accgttaaat ttgttgaatc cccattgcca 780ggtcaccaag gagaaaaaag aacgtattac cttccagcgc aacctgtaaa tgaagatgat 840ctgatttctt ctgtatttag taacattcca attattaatc ctgtttctaa tctataatga 90026298PRTBacillus thuringiensis 26Met Ala Ile Phe Asn Phe Asp Ala Lys Val Val Glu Phe Ile Asn Trp1 5 10 15Trp Thr Ala Glu Phe Gly Gly Lys Asp Pro Arg Asn Ile Gln Ile Gly 20 25 30Tyr Glu Asn Arg Asp Ile Asn Val Val Pro Ser Ser Ser Asn Pro Ser 35 40 45Val Asn Val Ile Pro Lys Leu Ala Arg Ser Ser Val Gln Glu Leu Gln 50 55 60Asn Asn Thr Ser Val Thr Gln Thr Gln Glu Leu Ala Phe Ser Glu Thr65 70 75 80Thr Thr Glu Ser Gln Ser Ser Thr Thr Thr His Gly Ala Ser Phe Ser 85 90 95Thr Thr Val Thr Ser Val Thr Gln Phe Thr Ala Glu Val Asn Phe Lys 100 105 110Ala Ile Gly Ser Ser Ile Glu Gln Thr Ile Gly Val Ser Met Thr Gly 115 120 125Asp Tyr Asn Tyr Ser Ser Ser Leu Thr Lys Thr Thr Glu Lys Ser Arg 130 135 140Ser Trp Thr Leu Thr Gln Pro Val Val Val Pro Pro Phe Ser Arg Val145 150 155 160Thr Cys Thr Leu Leu Ile Tyr Asn Ala Pro Phe Ser Val Pro Val Asp 165 170 175Leu Asn Cys Asn Val Phe Gly Thr Leu Gly Gly Asp Phe Leu Ala Ser 180 185 190Tyr Thr Tyr Thr Val Ile Ser Thr Gly Arg Thr Val Asn Thr Ser Ile 195 200 205Thr Ala Ser Gln Met Thr Leu Thr Ser Trp Pro Gly Lys Pro Ser Glu 210 215 220Ile Ile Gly Ile Ala Pro Lys His Gly Leu Ile Phe Lys Gly Thr Gly225 230 235 240Thr Gln Ala Ala Val His Gly Leu Tyr Ser Thr Val Lys Phe Val Glu 245 250 255Ser Pro Leu Pro Gly His Gln Gly Glu Lys Arg Thr Tyr Tyr Leu Pro 260 265 270Ala Gln Pro Val Asn Glu Asp Asp Leu Ile Ser Ser Val Phe Ser Asn 275 280 285Ile Pro Ile Ile Asn Pro Val Ser Asn Leu 290 29527897DNAArtificial sequenceSynthetic 27atggctatct tcaacttcga cgccaaggtg gtcgagttca

tcaactggtg gactgctgag 60ttcggcggca aagatccaag gaacatccag atcggctacg agaatcgtga catcaacgtg 120gtccctagct ctagcaaccc tagcgtgaat gtgatcccga agctcgctag gtccagcgta 180caggagcttc agaacaacac cagcgttacc cagactcagg agctggcctt ctccgaaacg 240acaaccgaat cccagtcctc tactaccact cacggcgctt cctttagcac taccgtgacc 300tcagtgaccc aattcaccgc cgaggtcaac ttcaaggcaa tcggtagctc tattgagcaa 360acaatcggcg tgagcatgac aggcgattac aactactcct cttcgttgac caagacaacc 420gagaaatcac gctcctggac gctgacgcag cctgttgtgg ttcctccctt ctctcgcgtc 480acgtgcaccc tcctaatcta caacgcgccc ttcagtgttc cagtcgatct caattgcaac 540gtcttcggaa cgcttggagg cgactttctg gcctcgtaca cttacaccgt gatctcaacg 600ggaagaactg tcaacacctc gattaccgcc tctcagatga ctctcaccag ttggcctggg 660aagcccagtg agatcattgg tatcgctccg aagcacggcc tcatcttcaa gggcactggt 720acacaggcgg ccgtccacgg gttgtactca acggtcaagt tcgttgaatc gccgctccct 780ggccatcagg gcgagaaacg gacctactat ctgccagcac aaccagtgaa cgaggacgat 840cttatttcct cggtgttctc caacattccg atcatcaacc cagtgtccaa tctgtga 89728298PRTArtificial sequenceSynthetic 28Met Ala Ile Phe Asn Phe Asp Ala Lys Val Val Glu Phe Ile Asn Trp1 5 10 15Trp Thr Ala Glu Phe Gly Gly Lys Asp Pro Arg Asn Ile Gln Ile Gly 20 25 30Tyr Glu Asn Arg Asp Ile Asn Val Val Pro Ser Ser Ser Asn Pro Ser 35 40 45Val Asn Val Ile Pro Lys Leu Ala Arg Ser Ser Val Gln Glu Leu Gln 50 55 60Asn Asn Thr Ser Val Thr Gln Thr Gln Glu Leu Ala Phe Ser Glu Thr65 70 75 80Thr Thr Glu Ser Gln Ser Ser Thr Thr Thr His Gly Ala Ser Phe Ser 85 90 95Thr Thr Val Thr Ser Val Thr Gln Phe Thr Ala Glu Val Asn Phe Lys 100 105 110Ala Ile Gly Ser Ser Ile Glu Gln Thr Ile Gly Val Ser Met Thr Gly 115 120 125Asp Tyr Asn Tyr Ser Ser Ser Leu Thr Lys Thr Thr Glu Lys Ser Arg 130 135 140Ser Trp Thr Leu Thr Gln Pro Val Val Val Pro Pro Phe Ser Arg Val145 150 155 160Thr Cys Thr Leu Leu Ile Tyr Asn Ala Pro Phe Ser Val Pro Val Asp 165 170 175Leu Asn Cys Asn Val Phe Gly Thr Leu Gly Gly Asp Phe Leu Ala Ser 180 185 190Tyr Thr Tyr Thr Val Ile Ser Thr Gly Arg Thr Val Asn Thr Ser Ile 195 200 205Thr Ala Ser Gln Met Thr Leu Thr Ser Trp Pro Gly Lys Pro Ser Glu 210 215 220Ile Ile Gly Ile Ala Pro Lys His Gly Leu Ile Phe Lys Gly Thr Gly225 230 235 240Thr Gln Ala Ala Val His Gly Leu Tyr Ser Thr Val Lys Phe Val Glu 245 250 255Ser Pro Leu Pro Gly His Gln Gly Glu Lys Arg Thr Tyr Tyr Leu Pro 260 265 270Ala Gln Pro Val Asn Glu Asp Asp Leu Ile Ser Ser Val Phe Ser Asn 275 280 285Ile Pro Ile Ile Asn Pro Val Ser Asn Leu 290 29529807DNABacillus thuringiensis 29atgggaatta tcaacattca agacgaaatt aatgactaca tgaaaggtat gtatggtgca 60acatctgtta aaagcacata tgacccctca ttcaaagtat ttaacgaatc tgtaacacct 120caatatgatg tgatttcaac agaacctgta aataatcata ttactactaa agcaataaat 180aatccagcgt cttcagaagt aaccagtaca gtaaccttca catggacgga aaccgacact 240gtaacctctg cagtgactaa agggtataaa gtcggtggtt cagtaagctc aaaagcaact 300tttaaatttg cttttgttac ttctgatgtt actgtaactg tatcagcaga atataattat 360agtacaacag atacaacaac aaaaacagat acacgcacat ggacggattc gacgacagta 420aaagcccctc caagaactaa tgtagaagtt gcatatatta tccaaactgg aaattataac 480gttccggtta atgtagagtc tgatatgact ggaacgctat tttgcagagg gtatagagat 540ggtgcactaa ttgcagcgac ttatatttct ataacagatt tagcagatta caagcctaat 600ttgggtctta caaataaagg ggatggggtt gctcatttta aaggtgaagg ttatatagag 660ggtgcacaag gcttaagaag ctatattcaa gttacagaat atccaatgga tgataaagac 720agacgttcga caccaaaaac ttatacaatt gaaggttcat tagcacccaa tgttacttta 780ataaatgata gaaaggaagg tagataa 80730268PRTBacillus thuringiensis 30Met Gly Ile Ile Asn Ile Gln Asp Glu Ile Asn Asp Tyr Met Lys Gly1 5 10 15Met Tyr Gly Ala Thr Ser Val Lys Ser Thr Tyr Asp Pro Ser Phe Lys 20 25 30 Val Phe Asn Glu Ser Val Thr Pro Gln Tyr Asp Val Ile Ser Thr Glu 35 40 45Pro Val Asn Asn His Ile Thr Thr Lys Ala Ile Asn Asn Pro Ala Ser 50 55 60Ser Glu Val Thr Ser Thr Val Thr Phe Thr Trp Thr Glu Thr Asp Thr65 70 75 80Val Thr Ser Ala Val Thr Lys Gly Tyr Lys Val Gly Gly Ser Val Ser 85 90 95Ser Lys Ala Thr Phe Lys Phe Ala Phe Val Thr Ser Asp Val Thr Val 100 105 110Thr Val Ser Ala Glu Tyr Asn Tyr Ser Thr Thr Asp Thr Thr Thr Lys 115 120 125Thr Asp Thr Arg Thr Trp Thr Asp Ser Thr Thr Val Lys Ala Pro Pro 130 135 140Arg Thr Asn Val Glu Val Ala Tyr Ile Ile Gln Thr Gly Asn Tyr Asn145 150 155 160Val Pro Val Asn Val Glu Ser Asp Met Thr Gly Thr Leu Phe Cys Arg 165 170 175Gly Tyr Arg Asp Gly Ala Leu Ile Ala Ala Thr Tyr Ile Ser Ile Thr 180 185 190Asp Leu Ala Asp Tyr Lys Pro Asn Leu Gly Leu Thr Asn Lys Gly Asp 195 200 205Gly Val Ala His Phe Lys Gly Glu Gly Tyr Ile Glu Gly Ala Gln Gly 210 215 220Leu Arg Ser Tyr Ile Gln Val Thr Glu Tyr Pro Met Asp Asp Lys Asp225 230 235 240Arg Arg Ser Thr Pro Lys Thr Tyr Thr Ile Glu Gly Ser Leu Ala Pro 245 250 255Asn Val Thr Leu Ile Asn Asp Arg Lys Glu Gly Arg 260 26531807DNAArtificial sequenceSynthetic 31atgggcatta tcaacattca agatgagatt aacgattaca tgaagggtat gtacggtgct 60accagtgtga agtccactta cgatcctagt ttcaaggtct tcaacgagtc tgtgactcct 120caatacgatg tgatttctac tgagcctgtg aacaatcaca ttactactaa ggctattaac 180aatcctgcta gttctgaggt tactagcact gttactttca cttggactga gactgatact 240gttactagcg ctgttactaa gggttacaag gttggtggat ctgtttcttc aaaggctacc 300ttcaagtttg ctttcgttac ctcagatgtt accgttaccg tgagcgctga gtacaactac 360tcaaccacag ataccaccac caagaccgat accagaacct ggaccgattc aaccacagtg 420aaggcaccac ctcgcaccaa cgtggaggtc gcctacatta tccaaacagg caactacaat 480gtcccagtca atgtcgaatc cgacatgaca ggaacactct tctgccgtgg ctacagggat 540ggagcactta ttgcagccac ctacatctcc atcacagacc ttgccgacta caagccaaat 600ctcggactca caaacaaggg agacggcgtc gcccacttca aaggcgaagg ctacatcgaa 660ggtgcccagg gtctaaggag ctacatccaa gtgacagaat acccaatgga cgacaaagac 720cgtcgctcca cacccaagac gtacaccatc gaagggtcgt tagcgccgaa tgtaacgctg 780ataaatgacc gaaaggaagg gcggtga 80732268PRTArtificial sequenceSynthetic 32Met Gly Ile Ile Asn Ile Gln Asp Glu Ile Asn Asp Tyr Met Lys Gly1 5 10 15Met Tyr Gly Ala Thr Ser Val Lys Ser Thr Tyr Asp Pro Ser Phe Lys 20 25 30Val Phe Asn Glu Ser Val Thr Pro Gln Tyr Asp Val Ile Ser Thr Glu 35 40 45Pro Val Asn Asn His Ile Thr Thr Lys Ala Ile Asn Asn Pro Ala Ser 50 55 60Ser Glu Val Thr Ser Thr Val Thr Phe Thr Trp Thr Glu Thr Asp Thr65 70 75 80Val Thr Ser Ala Val Thr Lys Gly Tyr Lys Val Gly Gly Ser Val Ser 85 90 95Ser Lys Ala Thr Phe Lys Phe Ala Phe Val Thr Ser Asp Val Thr Val 100 105 110Thr Val Ser Ala Glu Tyr Asn Tyr Ser Thr Thr Asp Thr Thr Thr Lys 115 120 125Thr Asp Thr Arg Thr Trp Thr Asp Ser Thr Thr Val Lys Ala Pro Pro 130 135 140Arg Thr Asn Val Glu Val Ala Tyr Ile Ile Gln Thr Gly Asn Tyr Asn145 150 155 160Val Pro Val Asn Val Glu Ser Asp Met Thr Gly Thr Leu Phe Cys Arg 165 170 175 Gly Tyr Arg Asp Gly Ala Leu Ile Ala Ala Thr Tyr Ile Ser Ile Thr 180 185 190Asp Leu Ala Asp Tyr Lys Pro Asn Leu Gly Leu Thr Asn Lys Gly Asp 195 200 205Gly Val Ala His Phe Lys Gly Glu Gly Tyr Ile Glu Gly Ala Gln Gly 210 215 220Leu Arg Ser Tyr Ile Gln Val Thr Glu Tyr Pro Met Asp Asp Lys Asp225 230 235 240Arg Arg Ser Thr Pro Lys Thr Tyr Thr Ile Glu Gly Ser Leu Ala Pro 245 250 255Asn Val Thr Leu Ile Asn Asp Arg Lys Glu Gly Arg 260 26533381DNABacillus thuringiensis 33atgacagtat ataacgtaac ttttaccatt aaattcttta atcacggtga atgggggggg 60ccagaacctt acggtaagat atatgcatat cttcaaaatc cagatcataa tttcgaaatt 120tggtcacaag ataattgggg gaaggatacg cctgagaaaa gttctcacac tcaaacaatt 180aaaataagta gcccaacagg ggggcctata gaccaaatgt gtttttatgg tgatgtaaaa 240gaattcgacg taggaaattc agatgatgtt ctcgcctctc caagtcaaaa agtatgcagt 300acgcctggca caacaataag gcttaacgga gatgagagtg gttcttatat agagattagt 360tattccttgg ccccagctta a 38134126PRTBacillus thuringiensis 34Met Thr Val Tyr Asn Val Thr Phe Thr Ile Lys Phe Phe Asn His Gly1 5 10 15Glu Trp Gly Gly Pro Glu Pro Tyr Gly Lys Ile Tyr Ala Tyr Leu Gln 20 25 30Asn Pro Asp His Asn Phe Glu Ile Trp Ser Gln Asp Asn Trp Gly Lys 35 40 45Asp Thr Pro Glu Lys Ser Ser His Thr Gln Thr Ile Lys Ile Ser Ser 50 55 60Pro Thr Gly Gly Pro Ile Asp Gln Met Cys Phe Tyr Gly Asp Val Lys65 70 75 80Glu Phe Asp Val Gly Asn Ser Asp Asp Val Leu Ala Ser Pro Ser Gln 85 90 95Lys Val Cys Ser Thr Pro Gly Thr Thr Ile Arg Leu Asn Gly Asp Glu 100 105 110Ser Gly Ser Tyr Ile Glu Ile Ser Tyr Ser Leu Ala Pro Ala 115 120 12535381DNAArtificial sequenceSynthetic 35atgaccgtgt acaacgtgac cttcaccatt aagttcttca atcacggtga gtggggcggt 60cctgagccct acggcaaaat ctacgcctat ctccagaacc ctgaccacaa cttcgaaatc 120tggtcccagg acaactgggg caaggacact cccgagaagt ccagccatac gcagaccatc 180aagatcagca gcccgactgg cggcccgatt gaccagatgt gcttctacgg cgacgtcaag 240gaatttgacg tgggaaactc agatgacgtc ctcgccagcc cgtcccagaa ggtttgctcg 300acgccaggaa ctaccatccg cctgaatggc gatgaatctg ggtcgtacat cgagatcagt 360tacagccttg cgcctgcatg a 38136126PRTArtificial sequenceSynthetic 36Met Thr Val Tyr Asn Val Thr Phe Thr Ile Lys Phe Phe Asn His Gly1 5 10 15Glu Trp Gly Gly Pro Glu Pro Tyr Gly Lys Ile Tyr Ala Tyr Leu Gln 20 25 30Asn Pro Asp His Asn Phe Glu Ile Trp Ser Gln Asp Asn Trp Gly Lys 35 40 45Asp Thr Pro Glu Lys Ser Ser His Thr Gln Thr Ile Lys Ile Ser Ser 50 55 60Pro Thr Gly Gly Pro Ile Asp Gln Met Cys Phe Tyr Gly Asp Val Lys65 70 75 80Glu Phe Asp Val Gly Asn Ser Asp Asp Val Leu Ala Ser Pro Ser Gln 85 90 95Lys Val Cys Ser Thr Pro Gly Thr Thr Ile Arg Leu Asn Gly Asp Glu 100 105 110Ser Gly Ser Tyr Ile Glu Ile Ser Tyr Ser Leu Ala Pro Ala 115 120 12537906DNABacillus thuringiensis 37atgaacaata attctaaatt tttattctca cctataaatg ctaatggaaa taatccttat 60tttttagatg tagataattg gaaagatgta ttttgttatc ttgtaaatac tctggaaatg 120aaatatacaa aagtagttaa taaatctcaa ttaatagatg cattatttcc agatcctaat 180gatcaaagaa taggtccatt ttattggaat atatatgata attataaatt agataatgta 240ttaaaaccat tatatgattt tcatagttca ttattaggaa atattcttga atggggatta 300tttggaacaa taagaatggg tagtttggct tatcatattc aagattttgt aaatatcata 360tgtgtaaaac atataaatga atttaatcaa attatagatg atttacaaca agataatgtt 420tcaacaaaaa ctgttgataa atttagaaat atatgtattg aactacgtac taaatcactt 480ctttttcctc aagaagtaaa taaaatatta gatgcaagta gaaatcttgc aaataatctt 540ataaaagtta atgctatttt agattcaaca gtgcaacaat ttgaaaacgt ttatcaatca 600tcacaatcaa ataacgataa atattatttg agaatcgtaa aagaatattt aggaaacgaa 660caacaagatt tagaaaaaca aactcctaat catcttaata ctcttcaaaa attacgtgga 720atatggactg tttttggaga taatttagaa aaattaattc atgcttgtga tgatgatctt 780atagattttg atgcaatgat agctagtata aatttagagg atgctataac ttcttggaaa 840gaagtaaaaa ataatataga taaatttata ccagaatggg aaatctttaa aaaacaagga 900tgctaa 90638301PRTBacillus thuringiensis 38Met Asn Asn Asn Ser Lys Phe Leu Phe Ser Pro Ile Asn Ala Asn Gly1 5 10 15Asn Asn Pro Tyr Phe Leu Asp Val Asp Asn Trp Lys Asp Val Phe Cys 20 25 30Tyr Leu Val Asn Thr Leu Glu Met Lys Tyr Thr Lys Val Val Asn Lys 35 40 45Ser Gln Leu Ile Asp Ala Leu Phe Pro Asp Pro Asn Asp Gln Arg Ile 50 55 60Gly Pro Phe Tyr Trp Asn Ile Tyr Asp Asn Tyr Lys Leu Asp Asn Val65 70 75 80Leu Lys Pro Leu Tyr Asp Phe His Ser Ser Leu Leu Gly Asn Ile Leu 85 90 95Glu Trp Gly Leu Phe Gly Thr Ile Arg Met Gly Ser Leu Ala Tyr His 100 105 110Ile Gln Asp Phe Val Asn Ile Ile Cys Val Lys His Ile Asn Glu Phe 115 120 125Asn Gln Ile Ile Asp Asp Leu Gln Gln Asp Asn Val Ser Thr Lys Thr 130 135 140Val Asp Lys Phe Arg Asn Ile Cys Ile Glu Leu Arg Thr Lys Ser Leu145 150 155 160Leu Phe Pro Gln Glu Val Asn Lys Ile Leu Asp Ala Ser Arg Asn Leu 165 170 175Ala Asn Asn Leu Ile Lys Val Asn Ala Ile Leu Asp Ser Thr Val Gln 180 185 190Gln Phe Glu Asn Val Tyr Gln Ser Ser Gln Ser Asn Asn Asp Lys Tyr 195 200 205Tyr Leu Arg Ile Val Lys Glu Tyr Leu Gly Asn Glu Gln Gln Asp Leu 210 215 220Glu Lys Gln Thr Pro Asn His Leu Asn Thr Leu Gln Lys Leu Arg Gly225 230 235 240Ile Trp Thr Val Phe Gly Asp Asn Leu Glu Lys Leu Ile His Ala Cys 245 250 255 Asp Asp Asp Leu Ile Asp Phe Asp Ala Met Ile Ala Ser Ile Asn Leu 260 265 270Glu Asp Ala Ile Thr Ser Trp Lys Glu Val Lys Asn Asn Ile Asp Lys 275 280 285Phe Ile Pro Glu Trp Glu Ile Phe Lys Lys Gln Gly Cys 290 295 30039906DNAArtificial sequenceSynthetic 39atgaacaaca actctaagtt cctcttctcg cccatcaacg ctaacggcaa caacccttac 60ttcctagacg tggacaactg gaaggacgtc ttctgctacc tagtgaacac ccttgagatg 120aagtacacta aggtggtgaa caagtcccag ctcattgacg ccctcttccc agatcctaac 180gatcagcgta tcggaccctt ctactggaac atctacgata actacaagct cgacaacgtc 240ctcaagccac tgtacgactt tcacagctca ctcctgggca acatccttga gtggggactc 300tttgggacca ttcgcatggg ttctctggcc taccacatcc aagatttcgt caacattatc 360tgcgttaagc acatcaacga gttcaaccag ataattgatg acctccagca agataacgtc 420agcactaaga cggtggacaa gttccgcaac atctgcatcg aactccgcac gaagtccctt 480ctgttccctc aggaggtcaa taagattctg gatgcgtcac ggaacctcgc aaataacctc 540atcaaggtta atgcaatcct cgatagtacc gtccaacagt tcgagaacgt ttaccagtcg 600tctcagtcta ataacgacaa gtactatttg agaatcgtaa aggagtatct cgggaatgag 660cagcaagacc tggagaagca gacaccgaat cacctgaaca cccttcagaa gctccgtggc 720atctggacag tcttcggcga caatcttgag aagctcattc atgcttgtga tgacgatctc 780atcgacttcg acgctatgat cgcctccatc aacttggaag atgcgatcac ttcgtggaag 840gaggttaaga ataacattga caaattcatc cctgaatggg aaatcttcaa gaaacagggc 900tgctga 90640301PRTArtificial sequenceSynthetic 40Met Asn Asn Asn Ser Lys Phe Leu Phe Ser Pro Ile Asn Ala Asn Gly1 5 10 15Asn Asn Pro Tyr Phe Leu Asp Val Asp Asn Trp Lys Asp Val Phe Cys 20 25 30 Tyr Leu Val Asn Thr Leu Glu Met Lys Tyr Thr Lys Val Val Asn Lys 35 40 45Ser Gln Leu Ile Asp Ala Leu Phe Pro Asp Pro Asn Asp Gln Arg Ile 50 55 60Gly Pro Phe Tyr Trp Asn Ile Tyr Asp Asn Tyr Lys Leu Asp Asn Val65 70 75 80Leu Lys Pro Leu Tyr Asp Phe His Ser Ser Leu Leu Gly Asn Ile Leu 85 90 95Glu Trp Gly Leu Phe Gly Thr Ile Arg Met Gly Ser Leu Ala Tyr His 100 105 110Ile Gln Asp Phe Val Asn Ile Ile Cys Val Lys His Ile Asn Glu Phe 115 120 125Asn Gln Ile Ile Asp Asp Leu Gln Gln Asp Asn Val Ser Thr Lys Thr 130 135 140Val Asp Lys Phe Arg Asn Ile Cys Ile Glu Leu Arg Thr Lys Ser Leu145

150 155 160Leu Phe Pro Gln Glu Val Asn Lys Ile Leu Asp Ala Ser Arg Asn Leu 165 170 175Ala Asn Asn Leu Ile Lys Val Asn Ala Ile Leu Asp Ser Thr Val Gln 180 185 190Gln Phe Glu Asn Val Tyr Gln Ser Ser Gln Ser Asn Asn Asp Lys Tyr 195 200 205Tyr Leu Arg Ile Val Lys Glu Tyr Leu Gly Asn Glu Gln Gln Asp Leu 210 215 220Glu Lys Gln Thr Pro Asn His Leu Asn Thr Leu Gln Lys Leu Arg Gly225 230 235 240Ile Trp Thr Val Phe Gly Asp Asn Leu Glu Lys Leu Ile His Ala Cys 245 250 255Asp Asp Asp Leu Ile Asp Phe Asp Ala Met Ile Ala Ser Ile Asn Leu 260 265 270Glu Asp Ala Ile Thr Ser Trp Lys Glu Val Lys Asn Asn Ile Asp Lys 275 280 285Phe Ile Pro Glu Trp Glu Ile Phe Lys Lys Gln Gly Cys 290 295 30041861DNABacillus thuringiensis 41atgaataata ttaataagaa gtatagactt aatgatatga ttaataaaaa tcaatttatt 60atttcaaaaa cagaatgggt tactataaga acatatattg aaattggatt aactttacca 120gtaaatgaac aagatttacg aaaatatttc aatttaaatc cagatataac actatctaat 180gatttttctg aattatttga tatttgttat tctattaaaa atttagctca atggtggaat 240accactatac ttcctttaat tattaaatct gttaataata ttacatcata tggatttaaa 300attgctggta atccttttaa taataaagaa ggatactttt caaaattaca aaatgaatta 360catattatta ataattataa ttctaataaa acaacaaaaa ctattaaaca atttcaatct 420cggtgtaaca ttttaattaa ggaagttaaa caatatgaag atgttaccaa aaatattgta 480atattattaa ataaactttt atatggtaat cagcaaaaat tagaagggat tattaatatt 540caaaaacgat taaaagtggt tcaaacaact tttaatccga tatctaatga aactaattta 600atttataaaa aactatttga aaaaataaaa aaaataaata ttggtttttt tgaatgcgta 660aataaatata taagtatatt taacaaaata attattatgt ggtcaaatac tgaacaacaa 720attatagatt ttaaatcaat tttatttcaa gaatttaaaa atataaatga aacagttatt 780gaatttgaag atattattga gatttggtta attatagcta aaaaatctcg tgaatttact 840ttaaatgctt atatatctta g 86142286PRTBacillus thuringiensis 42Met Asn Asn Ile Asn Lys Lys Tyr Arg Leu Asn Asp Met Ile Asn Lys1 5 10 15Asn Gln Phe Ile Ile Ser Lys Thr Glu Trp Val Thr Ile Arg Thr Tyr 20 25 30Ile Glu Ile Gly Leu Thr Leu Pro Val Asn Glu Gln Asp Leu Arg Lys 35 40 45Tyr Phe Asn Leu Asn Pro Asp Ile Thr Leu Ser Asn Asp Phe Ser Glu 50 55 60Leu Phe Asp Ile Cys Tyr Ser Ile Lys Asn Leu Ala Gln Trp Trp Asn65 70 75 80Thr Thr Ile Leu Pro Leu Ile Ile Lys Ser Val Asn Asn Ile Thr Ser 85 90 95Tyr Gly Phe Lys Ile Ala Gly Asn Pro Phe Asn Asn Lys Glu Gly Tyr 100 105 110Phe Ser Lys Leu Gln Asn Glu Leu His Ile Ile Asn Asn Tyr Asn Ser 115 120 125Asn Lys Thr Thr Lys Thr Ile Lys Gln Phe Gln Ser Arg Cys Asn Ile 130 135 140Leu Ile Lys Glu Val Lys Gln Tyr Glu Asp Val Thr Lys Asn Ile Val145 150 155 160Ile Leu Leu Asn Lys Leu Leu Tyr Gly Asn Gln Gln Lys Leu Glu Gly 165 170 175Ile Ile Asn Ile Gln Lys Arg Leu Lys Val Val Gln Thr Thr Phe Asn 180 185 190Pro Ile Ser Asn Glu Thr Asn Leu Ile Tyr Lys Lys Leu Phe Glu Lys 195 200 205Ile Lys Lys Ile Asn Ile Gly Phe Phe Glu Cys Val Asn Lys Tyr Ile 210 215 220Ser Ile Phe Asn Lys Ile Ile Ile Met Trp Ser Asn Thr Glu Gln Gln225 230 235 240Ile Ile Asp Phe Lys Ser Ile Leu Phe Gln Glu Phe Lys Asn Ile Asn 245 250 255Glu Thr Val Ile Glu Phe Glu Asp Ile Ile Glu Ile Trp Leu Ile Ile 260 265 270Ala Lys Lys Ser Arg Glu Phe Thr Leu Asn Ala Tyr Ile Ser 275 280 28543861DNAArtificial sequenceSynthetic 43atgaacaaca ttaacaagaa gtaccgcctg aacgacatga ttaacaagaa ccagttcatc 60attagcaaga ctgagtgggt gaccatcagg acgtacatcg agatcggcct gaccctgccc 120gtgaacgagc aagacctgag gaagtacttc aacctcaacc ctgacattac actgagcaac 180gacttctcgg agctgttcga catctgctac tctatcaaga atctcgccca atggtggaac 240acaacgatcc ttccgctaat cattaagtct gtgaacaaca tcactagcta tggcttcaag 300atcgcgggca acccgttcaa caataaggag ggttacttta gtaagctcca gaacgaactc 360cacatcatta acaattacaa ttcgaacaag acgactaaga cgatcaagca gtttcagtca 420cggtgcaaca tcttgattaa ggaggttaaa caatacgaag atgtaacgaa gaacatcgtg 480atcctcctta acaagctcct gtacggcaac cagcagaagc tcgaagggat cattaacata 540cagaagcgtc tcaaggtggt ccagaccacg ttcaatccca tctccaatga gactaacctt 600atctacaaga aactctttga gaagatcaag aagatcaaca tcggcttctt tgagtgtgtc 660aacaagtaca tctccatctt caacaagata atcattatgt ggtcaaacac tgagcagcag 720atcatcgact tcaagtccat actgttccag gagttcaaga acatcaatga gaccgtcatc 780gagttcgaag acatcatcga gatttggctt atcatcgcta agaagtctag ggagttcacc 840ctgaatgcgt acatctcctg a 86144286PRTArtificial sequenceSynthetic 44Met Asn Asn Ile Asn Lys Lys Tyr Arg Leu Asn Asp Met Ile Asn Lys1 5 10 15Asn Gln Phe Ile Ile Ser Lys Thr Glu Trp Val Thr Ile Arg Thr Tyr 20 25 30Ile Glu Ile Gly Leu Thr Leu Pro Val Asn Glu Gln Asp Leu Arg Lys 35 40 45Tyr Phe Asn Leu Asn Pro Asp Ile Thr Leu Ser Asn Asp Phe Ser Glu 50 55 60Leu Phe Asp Ile Cys Tyr Ser Ile Lys Asn Leu Ala Gln Trp Trp Asn65 70 75 80Thr Thr Ile Leu Pro Leu Ile Ile Lys Ser Val Asn Asn Ile Thr Ser 85 90 95Tyr Gly Phe Lys Ile Ala Gly Asn Pro Phe Asn Asn Lys Glu Gly Tyr 100 105 110Phe Ser Lys Leu Gln Asn Glu Leu His Ile Ile Asn Asn Tyr Asn Ser 115 120 125Asn Lys Thr Thr Lys Thr Ile Lys Gln Phe Gln Ser Arg Cys Asn Ile 130 135 140Leu Ile Lys Glu Val Lys Gln Tyr Glu Asp Val Thr Lys Asn Ile Val145 150 155 160Ile Leu Leu Asn Lys Leu Leu Tyr Gly Asn Gln Gln Lys Leu Glu Gly 165 170 175Ile Ile Asn Ile Gln Lys Arg Leu Lys Val Val Gln Thr Thr Phe Asn 180 185 190Pro Ile Ser Asn Glu Thr Asn Leu Ile Tyr Lys Lys Leu Phe Glu Lys 195 200 205Ile Lys Lys Ile Asn Ile Gly Phe Phe Glu Cys Val Asn Lys Tyr Ile 210 215 220Ser Ile Phe Asn Lys Ile Ile Ile Met Trp Ser Asn Thr Glu Gln Gln225 230 235 240Ile Ile Asp Phe Lys Ser Ile Leu Phe Gln Glu Phe Lys Asn Ile Asn 245 250 255Glu Thr Val Ile Glu Phe Glu Asp Ile Ile Glu Ile Trp Leu Ile Ile 260 265 270Ala Lys Lys Ser Arg Glu Phe Thr Leu Asn Ala Tyr Ile Ser 275 280 28545960DNABacillus thuringiensis 45atgacaatta caaatatcga attagctata cgagattata caaattggga tggtactcgt 60gaaattccgg gatacatcaa tcgtcaggtt atagatgggc caaatatata tgactatgta 120attagtgact ctgtagctgt tccaaaaact gtaattttca atgtaaatcc aactccatat 180accgggccta atataatatc agaaaataat acagatgtaa atcaaaataa acgtattaag 240ttttctgaaa aagtagttga aactactaca catactacta caaagggttt taaaattggc 300ggtggaatta aatctactac aaaaggaact ttaaaattaa aatttcctgt aggagaacta 360gggtttgagc aaactcttga gctacctctt acaggagaat acaatagtag ttccactacg 420gggaacactt gtgcaaatga aaaattatgg gaaataacag ataatataac tgtacctcca 480cattcacgtg taacttcaac tttaataatt atgaaaacgg aagtaagggt tccaatggaa 540ttaaccacta atcttagagg aactaattct agtggtgaag gctcgttccc tactagtaat 600ggtctttttt catatactac ttcagctcgt ggaactgtag gcggtatttt tgttagttat 660tacgtgaggc ctgcttctgc attgtataat acctcttggc ctgataaacc tgcaactttt 720aattctattg gctcaaatga atctctaaat ttattgggat ctggatattc tgacgtagtt 780ccatctctat atgttactat tagacaagat caaactccat tatcaggata tccaggtgaa 840acgaaaacct ggtattcaga taaagtgata ttaagagatg gaagaattgt aacactacca 900agcaatgctg atgtaaatat gtcacaaaca gccaaaattc catattgtga tagatcttaa 96046319PRTBacillus thuringiensis 46Met Thr Ile Thr Asn Ile Glu Leu Ala Ile Arg Asp Tyr Thr Asn Trp1 5 10 15Asp Gly Thr Arg Glu Ile Pro Gly Tyr Ile Asn Arg Gln Val Ile Asp 20 25 30Gly Pro Asn Ile Tyr Asp Tyr Val Ile Ser Asp Ser Val Ala Val Pro 35 40 45Lys Thr Val Ile Phe Asn Val Asn Pro Thr Pro Tyr Thr Gly Pro Asn 50 55 60Ile Ile Ser Glu Asn Asn Thr Asp Val Asn Gln Asn Lys Arg Ile Lys65 70 75 80Phe Ser Glu Lys Val Val Glu Thr Thr Thr His Thr Thr Thr Lys Gly 85 90 95Phe Lys Ile Gly Gly Gly Ile Lys Ser Thr Thr Lys Gly Thr Leu Lys 100 105 110Leu Lys Phe Pro Val Gly Glu Leu Gly Phe Glu Gln Thr Leu Glu Leu 115 120 125Pro Leu Thr Gly Glu Tyr Asn Ser Ser Ser Thr Thr Gly Asn Thr Cys 130 135 140Ala Asn Glu Lys Leu Trp Glu Ile Thr Asp Asn Ile Thr Val Pro Pro145 150 155 160His Ser Arg Val Thr Ser Thr Leu Ile Ile Met Lys Thr Glu Val Arg 165 170 175Val Pro Met Glu Leu Thr Thr Asn Leu Arg Gly Thr Asn Ser Ser Gly 180 185 190Glu Gly Ser Phe Pro Thr Ser Asn Gly Leu Phe Ser Tyr Thr Thr Ser 195 200 205Ala Arg Gly Thr Val Gly Gly Ile Phe Val Ser Tyr Tyr Val Arg Pro 210 215 220Ala Ser Ala Leu Tyr Asn Thr Ser Trp Pro Asp Lys Pro Ala Thr Phe225 230 235 240Asn Ser Ile Gly Ser Asn Glu Ser Leu Asn Leu Leu Gly Ser Gly Tyr 245 250 255Ser Asp Val Val Pro Ser Leu Tyr Val Thr Ile Arg Gln Asp Gln Thr 260 265 270Pro Leu Ser Gly Tyr Pro Gly Glu Thr Lys Thr Trp Tyr Ser Asp Lys 275 280 285Val Ile Leu Arg Asp Gly Arg Ile Val Thr Leu Pro Ser Asn Ala Asp 290 295 300Val Asn Met Ser Gln Thr Ala Lys Ile Pro Tyr Cys Asp Arg Ser305 310 31547960DNAArtificial sequenceSynthetic 47atgaccatca ccaacatcga actcgccatc cgcgactaca ctaactggga cggcactagg 60gagatccctg gctacatcaa ccgccaagtg atcgacggcc cgaacatcta cgactacgtg 120atctctgact ccgtggccgt gcctaagacc gtcatcttca acgtgaaccc aactccgtac 180actgggccaa acatcataag cgagaacaat actgacgtta atcagaacaa gcgcattaag 240ttctcggaga aggttgtgga gaccactacc catacgacca caaagggctt caagatcggt 300ggcggtatca agtccaccac gaagggcact ctcaagctga aattccctgt cggtgagctt 360ggatttgagc aaaccctcga acttcctctg acgggcgagt acaactccag ctccaccact 420gggaacacct gcgccaacga gaaactgtgg gaaattaccg acaacataac cgtcccgcca 480cactcccgag tcacctctac gcttatcatt atgaagacag aagtaagggt gccgatggag 540ttgacgacta acctgcgtgg aacaaacagt tccggcgaag ggagctttcc tacctctaat 600ggcctcttta gctacaccac atcagctaga ggaaccgttg gcggtatctt cgtttcttac 660tacgtccgtc cggccagcgc actctacaac acatcatggc ctgacaagcc cgcgaccttc 720aatagcattg gtagcaatga gtcgctcaac ctgctcggtt ccggttacag cgatgttgtg 780cccagtctct atgttacgat ccggcaagat cagactccgc tgtccggcta cccaggcgag 840acaaagacat ggtactccga taaggtcatc cttcgtgatg ggcgcattgt caccctccct 900tctaatgcgg atgtcaacat gagccagacg gctaagatcc cgtattgcga ccggtcctga 96048319PRTArtificial sequenceSynthetic 48Met Thr Ile Thr Asn Ile Glu Leu Ala Ile Arg Asp Tyr Thr Asn Trp1 5 10 15Asp Gly Thr Arg Glu Ile Pro Gly Tyr Ile Asn Arg Gln Val Ile Asp 20 25 30Gly Pro Asn Ile Tyr Asp Tyr Val Ile Ser Asp Ser Val Ala Val Pro 35 40 45Lys Thr Val Ile Phe Asn Val Asn Pro Thr Pro Tyr Thr Gly Pro Asn 50 55 60Ile Ile Ser Glu Asn Asn Thr Asp Val Asn Gln Asn Lys Arg Ile Lys65 70 75 80Phe Ser Glu Lys Val Val Glu Thr Thr Thr His Thr Thr Thr Lys Gly 85 90 95Phe Lys Ile Gly Gly Gly Ile Lys Ser Thr Thr Lys Gly Thr Leu Lys 100 105 110Leu Lys Phe Pro Val Gly Glu Leu Gly Phe Glu Gln Thr Leu Glu Leu 115 120 125Pro Leu Thr Gly Glu Tyr Asn Ser Ser Ser Thr Thr Gly Asn Thr Cys 130 135 140Ala Asn Glu Lys Leu Trp Glu Ile Thr Asp Asn Ile Thr Val Pro Pro145 150 155 160His Ser Arg Val Thr Ser Thr Leu Ile Ile Met Lys Thr Glu Val Arg 165 170 175Val Pro Met Glu Leu Thr Thr Asn Leu Arg Gly Thr Asn Ser Ser Gly 180 185 190Glu Gly Ser Phe Pro Thr Ser Asn Gly Leu Phe Ser Tyr Thr Thr Ser 195 200 205Ala Arg Gly Thr Val Gly Gly Ile Phe Val Ser Tyr Tyr Val Arg Pro 210 215 220Ala Ser Ala Leu Tyr Asn Thr Ser Trp Pro Asp Lys Pro Ala Thr Phe225 230 235 240Asn Ser Ile Gly Ser Asn Glu Ser Leu Asn Leu Leu Gly Ser Gly Tyr 245 250 255Ser Asp Val Val Pro Ser Leu Tyr Val Thr Ile Arg Gln Asp Gln Thr 260 265 270Pro Leu Ser Gly Tyr Pro Gly Glu Thr Lys Thr Trp Tyr Ser Asp Lys 275 280 285Val Ile Leu Arg Asp Gly Arg Ile Val Thr Leu Pro Ser Asn Ala Asp 290 295 300Val Asn Met Ser Gln Thr Ala Lys Ile Pro Tyr Cys Asp Arg Ser305 310 31549855DNABacillus thuringiensis 49atgaaatatc acaaaaaatt tcatgaaata gcttgggagt ttgctgaaaa atggactgaa 60caagaaggtt tggagttagc aaatgtcgat tatgtgaatc ctgctactgg taaagacacg 120ttgaatttcg tgaataaatt tgaatatatc ggaaaaataa aagagggaaa tctttgccca 180gagatagtag caaatgagtc tttttcaaat tcaaaatgtg atactttgaa aaataatctc 240cataaaaaat tatttttgaa acaatattat ttatgggata tagactatca atttataatt 300ccgccttcaa tttatacaaa tccaatatta ccgccatgtc ttagtaaaaa aatcaatcca 360gcaatacagg tagatttatt taaaagagca taccattttg aatctcaact aaacaattct 420gaaataatag aagcagggat ttatattgaa cctaatcaaa cgataaatgc taaagtaata 480gcagaatata aaaatgtgca acaaaaatat tgtatacacc ttaaaatttc aggaagtatt 540gttattgaag tgaaaaaaaa tcgtaattct tgtaaggatt ctaaaacatt ttatactatc 600ccaatcgtag atttgtataa atcagagctt gcacataatc attcttttca tttagatggg 660gagactgtta tatttactga aaaaggtatg tttaaaggcc taatttgttc taatgtattt 720atcgaagggg aacggtttaa tttaaaaaca ggagaatgct taggtaaata tataatacca 780ttaggtatgg acgaagaaaa agttctagaa aatagtaaat caatattttt taattcagaa 840aaaggaggaa tttaa 85550284PRTBacillus thuringiensis 50Met Lys Tyr His Lys Lys Phe His Glu Ile Ala Trp Glu Phe Ala Glu1 5 10 15Lys Trp Thr Glu Gln Glu Gly Leu Glu Leu Ala Asn Val Asp Tyr Val 20 25 30Asn Pro Ala Thr Gly Lys Asp Thr Leu Asn Phe Val Asn Lys Phe Glu 35 40 45Tyr Ile Gly Lys Ile Lys Glu Gly Asn Leu Cys Pro Glu Ile Val Ala 50 55 60Asn Glu Ser Phe Ser Asn Ser Lys Cys Asp Thr Leu Lys Asn Asn Leu65 70 75 80His Lys Lys Leu Phe Leu Lys Gln Tyr Tyr Leu Trp Asp Ile Asp Tyr 85 90 95Gln Phe Ile Ile Pro Pro Ser Ile Tyr Thr Asn Pro Ile Leu Pro Pro 100 105 110Cys Leu Ser Lys Lys Ile Asn Pro Ala Ile Gln Val Asp Leu Phe Lys 115 120 125Arg Ala Tyr His Phe Glu Ser Gln Leu Asn Asn Ser Glu Ile Ile Glu 130 135 140Ala Gly Ile Tyr Ile Glu Pro Asn Gln Thr Ile Asn Ala Lys Val Ile145 150 155 160Ala Glu Tyr Lys Asn Val Gln Gln Lys Tyr Cys Ile His Leu Lys Ile 165 170 175Ser Gly Ser Ile Val Ile Glu Val Lys Lys Asn Arg Asn Ser Cys Lys 180 185 190Asp Ser Lys Thr Phe Tyr Thr Ile Pro Ile Val Asp Leu Tyr Lys Ser 195 200 205Glu Leu Ala His Asn His Ser Phe His Leu Asp Gly Glu Thr Val Ile 210 215 220Phe Thr Glu Lys Gly Met Phe Lys Gly Leu Ile Cys Ser Asn Val Phe225 230 235 240Ile Glu Gly Glu Arg Phe Asn Leu Lys Thr Gly Glu Cys Leu Gly Lys 245 250 255Tyr Ile Ile Pro Leu Gly Met Asp Glu Glu Lys Val Leu Glu Asn Ser 260 265 270Lys Ser Ile Phe Phe Asn Ser Glu Lys Gly Gly Ile 275 28051855DNAArtificial sequenceSynthetic 51atgaagtacc acaagaagtt ccacgagatc gcctgggagt tcgctgagaa gtggactgag 60caggagggcc tagagctggc taacgtggac tacgtgaacc ctgccaccgg caaggacacc 120ctgaacttcg tgaacaagtt

cgagtacatc ggaaagatta aggaggggaa cctctgtccc 180gagattgttg ccaatgagtc tttcagcaat agcaagtgcg atacgctcaa gaataacctc 240cacaagaaac tgttcttgaa gcagtactac ctgtgggaca tcgactacca gttcatcatt 300ccaccgagca tctacaccaa ccctatcctc cctccatgcc tgtcgaagaa gatcaacccg 360gccatccaag tggacctctt caagcgcgct taccactttg agtcccagct taacaatagc 420gagatcatcg aggccggtat ctacattgaa cccaaccaga ccatcaatgc gaaggttatc 480gcggagtaca agaacgtgca acagaagtat tgcatccacc tcaagatcag tggtagtatt 540gtcatcgagg tcaagaagaa caggaactcg tgcaaggact caaagacgtt ctacactatc 600ccgatagtgg acctttacaa gtctgaactc gcccacaacc attcattcca tctcgatgga 660gagactgtaa tcttcaccga gaagggcatg tttaagggcc ttatttgctc caacgtcttc 720atcgaaggcg aacgcttcaa ccttaagaca ggagaatgcc tggggaagta catcatccct 780ttgggcatgg atgaggagaa ggttctggag aactccaagt ctatcttctt caactccgag 840aagggcggga tctga 85552284PRTArtificial sequenceSynthetic 52Met Lys Tyr His Lys Lys Phe His Glu Ile Ala Trp Glu Phe Ala Glu1 5 10 15Lys Trp Thr Glu Gln Glu Gly Leu Glu Leu Ala Asn Val Asp Tyr Val 20 25 30Asn Pro Ala Thr Gly Lys Asp Thr Leu Asn Phe Val Asn Lys Phe Glu 35 40 45Tyr Ile Gly Lys Ile Lys Glu Gly Asn Leu Cys Pro Glu Ile Val Ala 50 55 60Asn Glu Ser Phe Ser Asn Ser Lys Cys Asp Thr Leu Lys Asn Asn Leu65 70 75 80His Lys Lys Leu Phe Leu Lys Gln Tyr Tyr Leu Trp Asp Ile Asp Tyr 85 90 95Gln Phe Ile Ile Pro Pro Ser Ile Tyr Thr Asn Pro Ile Leu Pro Pro 100 105 110Cys Leu Ser Lys Lys Ile Asn Pro Ala Ile Gln Val Asp Leu Phe Lys 115 120 125Arg Ala Tyr His Phe Glu Ser Gln Leu Asn Asn Ser Glu Ile Ile Glu 130 135 140Ala Gly Ile Tyr Ile Glu Pro Asn Gln Thr Ile Asn Ala Lys Val Ile145 150 155 160Ala Glu Tyr Lys Asn Val Gln Gln Lys Tyr Cys Ile His Leu Lys Ile 165 170 175Ser Gly Ser Ile Val Ile Glu Val Lys Lys Asn Arg Asn Ser Cys Lys 180 185 190Asp Ser Lys Thr Phe Tyr Thr Ile Pro Ile Val Asp Leu Tyr Lys Ser 195 200 205Glu Leu Ala His Asn His Ser Phe His Leu Asp Gly Glu Thr Val Ile 210 215 220Phe Thr Glu Lys Gly Met Phe Lys Gly Leu Ile Cys Ser Asn Val Phe225 230 235 240Ile Glu Gly Glu Arg Phe Asn Leu Lys Thr Gly Glu Cys Leu Gly Lys 245 250 255Tyr Ile Ile Pro Leu Gly Met Asp Glu Glu Lys Val Leu Glu Asn Ser 260 265 270Lys Ser Ile Phe Phe Asn Ser Glu Lys Gly Gly Ile 275 28053903DNABacillus thuringiensis 53atgctagcat tcatattttc aggaggaagt agccaacttt gtacgtcaat acgtaatgaa 60tataataaca tattgcaagg gggggggatg gatgtatctt catccttccg tgtagatcct 120tctaaaataa atataaatga tttggaagtg acatatccag agttaaatat tcccgataac 180ataatagctt ctatagtatc taataatagc ttcgagaata gaggatttat aactaacgaa 240acttacgcta ccgctgaggt aaccagagca ttaaccgaaa caattactac agctactact 300agagggttta aattcacgca aggttttacc tatacaaata agattaacct cagaatacca 360attgccggtt cagaatcaac aatttccttt tcaacatctt ttgaacaaaa tatttcaacc 420actgaaacaa taacaaaaac tgaaacgata actatacttg tacctagaca aacggttaca 480gtaaggccta gaacaagaaa agttgttcaa ataagacttt atcaattacg aattcctaga 540gtattcaccg aaatatctgc ttatgtaaca ggtactctta gacaaccaat tcctcaagcg 600aatcctgatg tttatgctac tctggtaagt gtcaataatg catgtcctaa tgcatttgtt 660aaccgtgaca attttttgag aatagatcgc gaaaagcggg gcttagcgtt aagaggagaa 720ggtgaattta gtggaaatat agtttcatta gatttcctga ttacgactac tgaatatgat 780ttggatacaa atgctattat taatatagat aataccttgg gcagagcagc tattctagga 840agtgagcctt gtggaactat tgcttataca gaaccaattg acattattca cactgattgt 900taa 90354300PRTBacillus thuringiensis 54Met Leu Ala Phe Ile Phe Ser Gly Gly Ser Ser Gln Leu Cys Thr Ser1 5 10 15Ile Arg Asn Glu Tyr Asn Asn Ile Leu Gln Gly Gly Gly Met Asp Val 20 25 30Ser Ser Ser Phe Arg Val Asp Pro Ser Lys Ile Asn Ile Asn Asp Leu 35 40 45Glu Val Thr Tyr Pro Glu Leu Asn Ile Pro Asp Asn Ile Ile Ala Ser 50 55 60Ile Val Ser Asn Asn Ser Phe Glu Asn Arg Gly Phe Ile Thr Asn Glu65 70 75 80Thr Tyr Ala Thr Ala Glu Val Thr Arg Ala Leu Thr Glu Thr Ile Thr 85 90 95Thr Ala Thr Thr Arg Gly Phe Lys Phe Thr Gln Gly Phe Thr Tyr Thr 100 105 110Asn Lys Ile Asn Leu Arg Ile Pro Ile Ala Gly Ser Glu Ser Thr Ile 115 120 125Ser Phe Ser Thr Ser Phe Glu Gln Asn Ile Ser Thr Thr Glu Thr Ile 130 135 140Thr Lys Thr Glu Thr Ile Thr Ile Leu Val Pro Arg Gln Thr Val Thr145 150 155 160Val Arg Pro Arg Thr Arg Lys Val Val Gln Ile Arg Leu Tyr Gln Leu 165 170 175Arg Ile Pro Arg Val Phe Thr Glu Ile Ser Ala Tyr Val Thr Gly Thr 180 185 190Leu Arg Gln Pro Ile Pro Gln Ala Asn Pro Asp Val Tyr Ala Thr Leu 195 200 205Val Ser Val Asn Asn Ala Cys Pro Asn Ala Phe Val Asn Arg Asp Asn 210 215 220Phe Leu Arg Ile Asp Arg Glu Lys Arg Gly Leu Ala Leu Arg Gly Glu225 230 235 240Gly Glu Phe Ser Gly Asn Ile Val Ser Leu Asp Phe Leu Ile Thr Thr 245 250 255Thr Glu Tyr Asp Leu Asp Thr Asn Ala Ile Ile Asn Ile Asp Asn Thr 260 265 270Leu Gly Arg Ala Ala Ile Leu Gly Ser Glu Pro Cys Gly Thr Ile Ala 275 280 285Tyr Thr Glu Pro Ile Asp Ile Ile His Thr Asp Cys 290 295 30055903DNAArtificial sequenceSynthetic 55atgcttgcct tcatcttctc cggcggtagc tcccagctct gcactagcat ccgtaacgag 60tacaacaaca tcctccaggg cggtggcatg gacgtcagct ccagcttccg cgtggaccct 120agcaagatca acatcaacga ccttgaggtg acctatcctg agttgaacat tccagataac 180attatcgcta gtattgtttc caacaacagt ttcgagaaca ggggattcat cactaacgag 240acttacgcca cggccgaagt aacccgcgct cttactgaga cgattacgac tgccaccacg 300cgcggattca agtttaccca ggggttcacc tacaccaaca agatcaacct ccgcatccca 360atcgctggct ctgagtccac catctccttc tctacgtcgt tcgaacagaa catctcgacc 420actgagacca tcacaaagac cgagactatc accatccttg tgccaaggca gaccgtcacc 480gtcagacctc ggaccaggaa ggtcgtgcaa atccgtcttt atcagttgcg tattccgagg 540gtgttcaccg agatttcagc gtatgttact ggcactctcc ggcagccgat ccctcaagca 600aacccggacg tttacgctac ccttgtttcc gtcaacaatg cctgccccaa tgctttcgtg 660aaccgagaca acttccttcg catcgaccgg gagaagcgcg gtctcgcgct gcgtggcgag 720ggcgagttct ctggaaacat cgtctccctg gatttcctga ttactacaac ggagtacgat 780ctggatacaa acgctatcat caacattgac aacacgctcg ggcgcgcggc catacttggg 840tcggaaccct gcggtacaat agcatacaca gagcccatcg acatcattca cacagattgt 900tga 90356300PRTArtificial sequenceSynthetic 56Met Leu Ala Phe Ile Phe Ser Gly Gly Ser Ser Gln Leu Cys Thr Ser1 5 10 15Ile Arg Asn Glu Tyr Asn Asn Ile Leu Gln Gly Gly Gly Met Asp Val 20 25 30Ser Ser Ser Phe Arg Val Asp Pro Ser Lys Ile Asn Ile Asn Asp Leu 35 40 45Glu Val Thr Tyr Pro Glu Leu Asn Ile Pro Asp Asn Ile Ile Ala Ser 50 55 60 Ile Val Ser Asn Asn Ser Phe Glu Asn Arg Gly Phe Ile Thr Asn Glu65 70 75 80Thr Tyr Ala Thr Ala Glu Val Thr Arg Ala Leu Thr Glu Thr Ile Thr 85 90 95Thr Ala Thr Thr Arg Gly Phe Lys Phe Thr Gln Gly Phe Thr Tyr Thr 100 105 110Asn Lys Ile Asn Leu Arg Ile Pro Ile Ala Gly Ser Glu Ser Thr Ile 115 120 125Ser Phe Ser Thr Ser Phe Glu Gln Asn Ile Ser Thr Thr Glu Thr Ile 130 135 140Thr Lys Thr Glu Thr Ile Thr Ile Leu Val Pro Arg Gln Thr Val Thr145 150 155 160Val Arg Pro Arg Thr Arg Lys Val Val Gln Ile Arg Leu Tyr Gln Leu 165 170 175Arg Ile Pro Arg Val Phe Thr Glu Ile Ser Ala Tyr Val Thr Gly Thr 180 185 190Leu Arg Gln Pro Ile Pro Gln Ala Asn Pro Asp Val Tyr Ala Thr Leu 195 200 205Val Ser Val Asn Asn Ala Cys Pro Asn Ala Phe Val Asn Arg Asp Asn 210 215 220Phe Leu Arg Ile Asp Arg Glu Lys Arg Gly Leu Ala Leu Arg Gly Glu225 230 235 240Gly Glu Phe Ser Gly Asn Ile Val Ser Leu Asp Phe Leu Ile Thr Thr 245 250 255Thr Glu Tyr Asp Leu Asp Thr Asn Ala Ile Ile Asn Ile Asp Asn Thr 260 265 270Leu Gly Arg Ala Ala Ile Leu Gly Ser Glu Pro Cys Gly Thr Ile Ala 275 280 285Tyr Thr Glu Pro Ile Asp Ile Ile His Thr Asp Cys 290 295 30057597DNABacillus thuringiensis 57atgaatacgt atcataaaat atgttctggt ccttatcagt ataatcacaa aggtccctac 60acggaatgtg ggcctactcc taggagatcc aatcccgatg aagaacattc aactttaaaa 120gattcatacg gtgatcctgt cctctattac atgccctatt atatggatcc atatgaattt 180cctggtgaaa gatttgtgta cgaatggact tctgatggag atcgtggagt taaattagga 240acgtcttatg cgaaagttta tttgcacttt tggaaagcta atggttccga tagtgcttta 300cgtatcaata tattggatcg tgagggaaac ccagaaaatc agtacttatc taagaaacgt 360ggtactgacg aagtactatt atataatcat gaccctctag aatccgaatg ggttccggag 420aattcatctg atcctaattt ttcagtagga aactacttct cactccaaaa tgcttcggat 480catattattt ccaatgaaaa attcttatcc tatagcattc caggtgaatg gttaaccaca 540gttcagccta ctatgaattc aaaaacaatg tggcgtttaa ttccaacatg gaaataa 59758198PRTBacillus thuringiensis 58Met Asn Thr Tyr His Lys Ile Cys Ser Gly Pro Tyr Gln Tyr Asn His1 5 10 15Lys Gly Pro Tyr Thr Glu Cys Gly Pro Thr Pro Arg Arg Ser Asn Pro 20 25 30Asp Glu Glu His Ser Thr Leu Lys Asp Ser Tyr Gly Asp Pro Val Leu 35 40 45Tyr Tyr Met Pro Tyr Tyr Met Asp Pro Tyr Glu Phe Pro Gly Glu Arg 50 55 60Phe Val Tyr Glu Trp Thr Ser Asp Gly Asp Arg Gly Val Lys Leu Gly65 70 75 80Thr Ser Tyr Ala Lys Val Tyr Leu His Phe Trp Lys Ala Asn Gly Ser 85 90 95Asp Ser Ala Leu Arg Ile Asn Ile Leu Asp Arg Glu Gly Asn Pro Glu 100 105 110Asn Gln Tyr Leu Ser Lys Lys Arg Gly Thr Asp Glu Val Leu Leu Tyr 115 120 125Asn His Asp Pro Leu Glu Ser Glu Trp Val Pro Glu Asn Ser Ser Asp 130 135 140Pro Asn Phe Ser Val Gly Asn Tyr Phe Ser Leu Gln Asn Ala Ser Asp145 150 155 160His Ile Ile Ser Asn Glu Lys Phe Leu Ser Tyr Ser Ile Pro Gly Glu 165 170 175Trp Leu Thr Thr Val Gln Pro Thr Met Asn Ser Lys Thr Met Trp Arg 180 185 190Leu Ile Pro Thr Trp Lys 19559660DNABacillus thuringiensis 59atgaatacgc atcataaaat gtgttctagt ccttatcgat ataatcatag acataaggat 60tgcgattgtc atttatataa ccataacggt ccttatcagt ataatcacag aggtccctac 120acggaatgtg gacctactcc taggagatca aatcccgatg aagaacattc aactttaaaa 180gattcatacg gtgatccagt ccgctatagc atgccctatt atatggatcc atatgaattt 240cctggtgaaa gatttgtgta cgaatggact tctgatggag atcgtggagt taaattagga 300acgtcttatg cgagtgttta tttgtacttt tggaaaatta ataatggtac cgatagtgct 360ttacgtatca atatattgga tcgtgaggga aacccagaaa atcagtactt atctaagaaa 420cgtggtactg acgaagtact attatataat cgtgaccctc tagaatccga atgggttccg 480gagaattcat ctgatcctaa tttttcagta ggaaactact tctcactcca aagtgcttcg 540gatcatatta tttccaatga aaaattctta tcctatagca ttcccggtga atggttaacc 600acagttcagc ctactatgaa ttcaaaaaca atgtggcgtt taattccagc atggaaataa 66060219PRTBacillus thuringiensis 60Met Asn Thr His His Lys Met Cys Ser Ser Pro Tyr Arg Tyr Asn His1 5 10 15Arg His Lys Asp Cys Asp Cys His Leu Tyr Asn His Asn Gly Pro Tyr 20 25 30Gln Tyr Asn His Arg Gly Pro Tyr Thr Glu Cys Gly Pro Thr Pro Arg 35 40 45Arg Ser Asn Pro Asp Glu Glu His Ser Thr Leu Lys Asp Ser Tyr Gly 50 55 60Asp Pro Val Arg Tyr Ser Met Pro Tyr Tyr Met Asp Pro Tyr Glu Phe65 70 75 80Pro Gly Glu Arg Phe Val Tyr Glu Trp Thr Ser Asp Gly Asp Arg Gly 85 90 95Val Lys Leu Gly Thr Ser Tyr Ala Ser Val Tyr Leu Tyr Phe Trp Lys 100 105 110Ile Asn Asn Gly Thr Asp Ser Ala Leu Arg Ile Asn Ile Leu Asp Arg 115 120 125Glu Gly Asn Pro Glu Asn Gln Tyr Leu Ser Lys Lys Arg Gly Thr Asp 130 135 140Glu Val Leu Leu Tyr Asn Arg Asp Pro Leu Glu Ser Glu Trp Val Pro145 150 155 160Glu Asn Ser Ser Asp Pro Asn Phe Ser Val Gly Asn Tyr Phe Ser Leu 165 170 175Gln Ser Ala Ser Asp His Ile Ile Ser Asn Glu Lys Phe Leu Ser Tyr 180 185 190Ser Ile Pro Gly Glu Trp Leu Thr Thr Val Gln Pro Thr Met Asn Ser 195 200 205Lys Thr Met Trp Arg Leu Ile Pro Ala Trp Lys 210 21561597DNAArtificial sequenceSynthetic 61atgaacacct accacaagat ttgctccggc ccttaccagt acaaccacaa gggaccgtac 60actgagtgcg gccctacccc taggcgtagc aaccctgacg aggagcactc cacccttaag 120gactcctacg gtgacccagt gctctactac atgccgtact acatggaccc atacgagttc 180cctggcgaaa ggttcgtgta cgagtggacg tcagatggcg atagaggagt caagctcggt 240actagctatg cgaaagttta cctacacttc tggaaggcta acgggtctga ctccgccttg 300cggatcaaca ttctcgatag ggagggcaac cctgagaacc agtatctctc caagaagcgc 360ggcaccgatg aggttcttct gtacaaccac gacccacttg agtccgaatg ggttccagaa 420aacagctcgg acccgaactt cagcgtggga aactacttca gtctccagaa cgcatcagat 480cacatcatct cgaatgagaa gtttctctct tattctatcc caggcgagtg gctgactacc 540gtgcaaccta cgatgaatag taagacaatg tggaggctca ttccgacatg gaagtga 59762198PRTArtificial sequenceSynthetic 62Met Asn Thr Tyr His Lys Ile Cys Ser Gly Pro Tyr Gln Tyr Asn His1 5 10 15Lys Gly Pro Tyr Thr Glu Cys Gly Pro Thr Pro Arg Arg Ser Asn Pro 20 25 30Asp Glu Glu His Ser Thr Leu Lys Asp Ser Tyr Gly Asp Pro Val Leu 35 40 45Tyr Tyr Met Pro Tyr Tyr Met Asp Pro Tyr Glu Phe Pro Gly Glu Arg 50 55 60Phe Val Tyr Glu Trp Thr Ser Asp Gly Asp Arg Gly Val Lys Leu Gly65 70 75 80Thr Ser Tyr Ala Lys Val Tyr Leu His Phe Trp Lys Ala Asn Gly Ser 85 90 95Asp Ser Ala Leu Arg Ile Asn Ile Leu Asp Arg Glu Gly Asn Pro Glu 100 105 110Asn Gln Tyr Leu Ser Lys Lys Arg Gly Thr Asp Glu Val Leu Leu Tyr 115 120 125Asn His Asp Pro Leu Glu Ser Glu Trp Val Pro Glu Asn Ser Ser Asp 130 135 140Pro Asn Phe Ser Val Gly Asn Tyr Phe Ser Leu Gln Asn Ala Ser Asp145 150 155 160His Ile Ile Ser Asn Glu Lys Phe Leu Ser Tyr Ser Ile Pro Gly Glu 165 170 175Trp Leu Thr Thr Val Gln Pro Thr Met Asn Ser Lys Thr Met Trp Arg 180 185 190Leu Ile Pro Thr Trp Lys 19563660DNAArtificial sequenceSynthetic 63atgaacaccc accacaagat gtgctctagc ccttaccgct acaaccaccg ccacaaggac 60tgcgactgcc acctctacaa ccacaacgga ccctaccagt acaaccatag gggcccttac 120actgagtgtg gccctactcc taggcgtagc aacccggacg aggaacacag caccctcaag 180gactcctacg gcgaccctgt gagatactcc atgccttact acatggaccc ttacgagttc 240ccaggcgagc gcttcgtgta cgagtggacc tccgacggcg acagaggtgt gaagctcggc 300acctcctacg cctccgtgta cctctacttc tggaagatca acaatggtac tgattccgct 360cttcggatca acatcctgga tagggagggc aacccggaga accagtatct gagcaagaaa 420cgcggcactg atgaggtgct gctttacaac cgtgatccgc tggagagcga gtgggtgccg 480gagaactctt cggaccctaa cttcagtgtt ggaaattact tcagtctcca gtcagcgtca 540gatcacatca tctctaatga gaagttcctg tcctactcga tcccaggaga atggctcacg 600acagttcaac ccacgatgaa ctcgaagaca atgtggcgat tgataccagc atggaaatga 66064219PRTArtificial sequencean amino acid sequence translation of SEQ ID NO 63. 64Met Asn Thr His His Lys Met Cys Ser Ser Pro Tyr Arg Tyr Asn His1 5 10 15Arg His Lys Asp Cys Asp Cys His Leu Tyr Asn His Asn Gly Pro Tyr 20 25 30Gln Tyr Asn His Arg Gly Pro Tyr Thr Glu Cys Gly Pro Thr Pro Arg 35 40 45Arg Ser Asn Pro

Asp Glu Glu His Ser Thr Leu Lys Asp Ser Tyr Gly 50 55 60Asp Pro Val Arg Tyr Ser Met Pro Tyr Tyr Met Asp Pro Tyr Glu Phe65 70 75 80Pro Gly Glu Arg Phe Val Tyr Glu Trp Thr Ser Asp Gly Asp Arg Gly 85 90 95Val Lys Leu Gly Thr Ser Tyr Ala Ser Val Tyr Leu Tyr Phe Trp Lys 100 105 110Ile Asn Asn Gly Thr Asp Ser Ala Leu Arg Ile Asn Ile Leu Asp Arg 115 120 125Glu Gly Asn Pro Glu Asn Gln Tyr Leu Ser Lys Lys Arg Gly Thr Asp 130 135 140Glu Val Leu Leu Tyr Asn Arg Asp Pro Leu Glu Ser Glu Trp Val Pro145 150 155 160Glu Asn Ser Ser Asp Pro Asn Phe Ser Val Gly Asn Tyr Phe Ser Leu 165 170 175Gln Ser Ala Ser Asp His Ile Ile Ser Asn Glu Lys Phe Leu Ser Tyr 180 185 190Ser Ile Pro Gly Glu Trp Leu Thr Thr Val Gln Pro Thr Met Asn Ser 195 200 205Lys Thr Met Trp Arg Leu Ile Pro Ala Trp Lys 210 215

Claims

1. A polynucleotide molecule encoding a pesticidal polypeptide, a) wherein the pesticidal polypeptide is optionally selected from the group consisting of: i) a protein having the amino acid sequence as set forth in any one of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; ii) a protein exhibiting an amino acid segment that is from about 45% to about 99.9% identical to the amino acid sequence as set forth in any one of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; or iii) a fragment of the protein as set forth in i) or ii) exhibiting substantially equivalent (biologically functional equivalent) pesticidal activity to that of the protein set forth in i) or ii); b) wherein said polynucleotide molecule is provided in a recombinant expression cassette; c) wherein said polynucleotide molecule is provided in a transgenic or recombinant host cell; i) wherein the host cell is selected from the group consisting of an Agrobacterium, a Bacillus, an Escherichia, a Salmonella, a Pseudomonas, a Rhizobium, and a plant cell; ii) wherein said plant cell is selected from the group of plant cells consisting of a alfalfa, banana, barley, bean, broccoli, cabbage, canola, carrot, cassava, castor, cauliflower, celery, chickpea, Chinese cabbage, citrus, coconut, coffee, corn, clover, cotton, a cucurbit, cucumber, Douglas fir, eggplant, eucalyptus, flax, garlic, grape, hops, leek, lettuce, Loblolly pine, millets, melons, nut, oat, olive, onion, ornamental, palm, pasture grass, pea, peanut, pepper, pigeonpea, pine, potato, poplar, pumpkin, Radiata pine, radish, rapeseed, rice, rootstocks, rye, safflower, shrub, sorghum, Southern pine, soybean, spinach, squash, strawberry, sugar beet, sugarcane, sunflower, sweet corn, sweet gum, sweet potato, switchgrass, tea, tobacco, tomato, triticale, turf grass, watermelon, and a wheat plant cell; d) wherein the polynucleotide molecule is provided in a recombinant plant selected from the group set forth in c) i); and e) wherein the polynucleotide molecule is present in a part of a recombinant plant, said plant part being selected from the group consisting of a leaf, a stem a flower, a sepal, a fruit, a root, or a seed; and i) wherein a product is produced from said recombinant plant part and said product comprises a detectable amount of said polynucleotide molecule, said product being selected from the group consisting of oil, meal, lint, and seed.

2. A pesticidal polypeptide: a) selected from the group consisting of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; b) having an amino acid sequence exhibiting from about 45% to about 99.9% identity to any one of the proteins set forth in SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; and c) a pesticidal fragment of the polypeptide as set forth in a) and b) exhibiting substantially equivalent (biologically functional equivalent) pesticidal activity of the polypeptide as set forth in a) or b).

3. A method of controlling pesticidal infection of a plant comprising optionally providing a pesticidal polypeptide: a) selected from the group consisting of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; b) having an amino acid sequence exhibiting from about 45% to about 99.9% identity to any one of the proteins set forth in SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; c) a pesticidal fragment of the polypeptide as set forth in a) and b) exhibiting substantially equivalent (biologically functional equivalent) pesticidal activity of the polypeptide as set forth in a) or b); d) as set forth in any of a), b), or c) to a pest susceptible to the polypeptide, thereby controlling the pest; e) in a pesticidally effective amount to a plant pest susceptible to the pesticidal polypeptide, thereby controlling the pest; wherein the pest is an insect or a nematode; wherein said insect is selected from the group consisting of any of the insect orders Coleoptera, Diptera, Hymenoptera, Lepidoptera, Mallophaga, Homoptera, Hemiptera, Orthoptera, Thysanoptera, Dermaptera, Isoptera, Anoplura, Siphonaptera, and Trichoptera; wherein said nematode is a nematode selected from the group consisting of Acontylus, Anguina, Aorolaimus, Aphasmatylenchus, Aphelenchoides, Aphelenchus, Atalodera, Atylenchus, Bakernema, Belonolaimus, Brachydrorus, Bursaphelenchus, Cacopaurus, Caloosia, Carphodorus, Criconema, Criconemella, Cryphodera, Ditylenchus, Dolichodorus, Eutylenchus, Globodera, Gracilacus, Helicotylenchus, Hemicriconemoides, Hemicycliophora, Heterodera, Hirschmanniella, Histotylenchus, Hoplolaimus, Hoplotylus, Longidorus, Macrotrophurus, Meloidodera, Meloidogyne, Merlinius, Morulaimus, Nacobbus, Nothanguina, Nothotylenchus, Paralongidorus, Paratrichodorus, Paratrophurus, Paratylenchus, Peltamigratus, Pratylenchoides, Pratylenchus, Psilenchus, Radopholoides, Radopholus, Rhadinaphelenchus, Rototylenchus, Rotylenchoides, Rotylenchus, Sarisodera, Scutellonema, Sphaeronema, Subanguina, Telotylenchoides, Telotylenchus, Trichotylenchus, Trophonema, Trophotylenculus, Trophurus, Tylenchorhynchus, Tylenchulus, Tylenchus, Tylodorus, Xiphinema, and Zygotylenchus nematodes; wherein the pesticidally effective amount of said pesticidal polypeptide is provided in the diet of the pest; wherein the pesticidally effective amount of said pesticidal polypeptide is provided in a recombinant plant, plant part, or product of said plant or plant part; wherein the pesticidally effective amount of said pesticidal polypeptide is provided in a topical formulation to a plant, plant part or seed and the formulation comprises bacterial cells, spores, or parasporal crystals comprising said polypeptide; and wherein the bacterial cells, spores, or parasporal crystals are obtained from a Bacillus species.

4. A method of: a) detecting in a biological sample a polynucleotide encoding a pesticidal protein having an amino acid sequence exhibiting from about 45% to about 99.9% amino acid sequence identity to any one of the proteins set forth in SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, SEQ ID NO:60, SEQ ID NO:62, and SEQ ID NO:64, wherein said method comprises the steps of: i) selecting a pair of primers that, when used together in an amplification reaction with said polynucleotide, form an amplicon encoding all or a pesticidally effective part of said pesticidal protein; ii) producing said amplicon from said polynucleotide; iii) detecting said amplicon; and iv) generating DNA sequence information from said amplicon to confirm the amino acid sequence identity of said pesticidal protein; b) detecting in a biological sample a pesticidal protein selected from the group consisting of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60 comprising the steps of: i) contacting said sample with an antibody that specifically binds to said pesticidal protein, or a peptide or epitope derived therefrom; and ii) detecting the binding of said antibody to said pesticidal protein, peptide, or epitope derived therefrom; and c) producing a crop from transgenic seed comprising a polynucleotide molecule encoding said pesticidal protein, the pesticidal protein being selected from the group consisting of: i) any one of the sequences selected from the group consisting of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; ii) a pesticidal protein having an amino acid sequence exhibiting from about 45% to about 99.9% identity to the amino acid sequence as set forth in any one of the sequences selected from the group consisting of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; and iii) a pesticidal fragment of any protein set forth in a) and b); wherein said method further comprises the steps of A) planting said seed; B) producing a crop from plants grown from said seed; and C) harvesting said crop; and wherein at least about 50% of the crop seed comprise said polynucleotide.

5. A transgenic plant, plant cell, seed, or plant part resistant to nematode and/or lepidopteran insect infestation comprising the polynucleotide of claim 1.

6. A commodity product (or biological sample) comprising a plant, seed, or plant part as set forth in claim 5, and further optionally comprising a detectable amount of: a) a polypeptide comprising the amino acid composition of the protein set forth in any of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; or b) a polynucleotide encoding any of the polypeptides of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; wherein detection of the polypeptide of a) or the polynucleotide of b) in said commodity (or biological sample) is determinative of the presence of said transgenic plant, plant cell, seed, or plant part in said commodity (or biological sample).

7. A method of producing a crop from seed comprising a polynucleotide molecule optionally encoding: a) a pesticidal polypeptide exhibiting an amino acid sequence selected from the group consisting of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; b) a pesticidal polypeptide having an amino acid sequence exhibiting at least 95% amino acid sequence identity to the amino acid sequence selected from the group consisting of SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:10, SEQ ID NO:14, SEQ ID NO:18, SEQ ID NO:22, SEQ ID NO:26, SEQ ID NO:30, SEQ ID NO:34, SEQ ID NO:38, SEQ ID NO:42, SEQ ID NO:46, SEQ ID NO:50, SEQ ID NO:54, SEQ ID NO:58, and SEQ ID NO:60; or c) a pesticidal fragment of the polypeptide of (a) or (b); wherein said method comprises the steps of planting the seed, producing a crop from plants grown from the seed, and harvesting the crop; and wherein at least 50% of the crop comprises seed comprising the polynucleotide molecule.

8. A composition comprising the pesticidal polypeptide of claim 2 in an agriculturally-acceptable carrier, and further comprising a recombinant Bacillus thuringiensis cell extract, cell suspension, cell homogenate, cell lysate, cell supernatant, cell filtrate, or cell pellet; wherein said composition comprises a pest inhibitory amount of said polypeptide and formulated as a powder, dust, pellet, granule, spray, emulsion, colloid, or solution; wherein said composition is prepared by desiccation, lyophilization, homogenization, extraction, filtration, centrifugation, sedimentation, or concentration of a culture of recombinant Bacillus thuringiensis cells comprising said polypeptide and said composition comprises from about 1% to about 99% by weight of said polypeptide.

9. The commodity product (or biological sample) of claim 6, wherein the polypeptide is detectable to a level of at least about (i) one part per million parts of such commodity product (or biological sample), or (ii) one nanogram per gram fresh weight of such commodity product (or biological sample).

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